Construction of an immune-related LncRNA signature with prognostic significance for bladder cancer.

Bladder cancer (BLCA) is one of the most common urological cancer with increasing cases and deaths every year. In the present study, we aim to construct an immune-related prognostic lncRNA signature (IRPLS) in bladder cancer (BLCA) patients and explore its immunogenomic implications in pan-cancers. First, the immune-related differentially expressed lncRNAs (IRDELs) were identified by 'limma' R package and the score of IRPLS in every patient were evaluated by Cox regression. The dysregulation of IRDELs expression between cancer and para-cancer normal tissues was validated through RT-qPCR. Then, we further explore the biological functions of a novel lncRNA from IRPLS, RP11-89 in BLCA using CCK8 assay, Transwell assay and Apoptosis analysis, which indicated that RP11-89 was able to promote cell proliferation and invasive capacity while inhibits cell apoptosis in BLCA. In addition, we performed bioinformatic methods and RIP to investigate and validate the RP11-89/miR-27a-3p/PPARγ pathway in order to explore the mechanism. Next, CIBERSORT and ESTIMATE algorithm were used to evaluate abundance of tumour-infiltrating immune cells and scores of tumour environment elements in BLCA with different level of IRPLS risk scores. Finally, multiple bioinformatic methods were performed to show us the immune landscape of these four lncRNAs for pan-cancers. In conclusion, this study first constructed an immune-related prognostic lncRNA signature, which consists of RP11-89, PSORS1C3, LINC02672 and MIR100HG and might shed lights on novel targets for individualized immunotherapy for BLCA patients.

Construction of a robust prognostic model for adult adrenocortical carcinoma: Results from bioinformatics and real-world data.

This study aims to construct a robust prognostic model for adult adrenocortical carcinoma (ACC) by large-scale multiomics analysis and real-world data. The RPPA data, gene expression profiles and clinical information of adult ACC patients were obtained from The Cancer Proteome Atlas (TCPA), Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA). Integrated prognosis-related proteins (IPRPs) model was constructed. Immunohistochemistry was used to validate the prognostic value of the IPRPs model in Fudan University Shanghai Cancer Center (FUSCC) cohort. 76 ACC cases from TCGA and 22 ACC cases from GSE10927 in NCBI's GEO database with full data for clinical information and gene expression were utilized to validate the effectiveness of the IPRPs model. Higher FASN (P = .039), FIBRONECTIN (P < .001), TFRC (P < .001), TSC1 (P < .001) expression indicated significantly worse overall survival for adult ACC patients. Risk assessment suggested significantly a strong predictive capacity of IPRPs model for poor overall survival (P < .05). IPRPs model showed a little stronger ability for predicting prognosis than Ki-67 protein in FUSCC cohort (P = .003, HR = 3.947; P = .005, HR = 3.787). In external validation of IPRPs model using gene expression data, IPRPs model showed strong ability for predicting prognosis in TCGA cohort (P = .005, HR = 3.061) and it exhibited best ability for predicting prognosis in GSE10927 cohort (P = .0898, HR = 2.318). This research constructed IPRPs model for predicting adult ACC patients' prognosis using proteomic data, gene expression data and real-world data and this prognostic model showed stronger predictive value than other biomarkers (Ki-67, Beta-catenin, etc) in multi-cohorts.

Primary tumor surgery improves survival in non-metastatic primary urethral carcinoma patients: a large population-based investigation.

BACKGROUND: Primary urethral carcinoma (PUC) is a rare genitourinary malignancy with a relatively poor prognosis. The aim of this study was to examine the impact of surgery on survival of patients diagnosed with PUC. METHODS: A total of 1544 PUC patients diagnosed between 2004 and 2016 were identified based on the SEER database. The Kaplan-Meier estimate and the Fine and Gray competing risks analysis were performed to assess overall survival (OS) and cancer-specific mortality (CSM). The multivariate Cox regression model and competing risks regression model were used to identify independent risk factors of OS and cancer-specific survival (CSS). RESULTS: The 5-yr OS was significantly better in patients who received either local therapy (39.8%) or radical surgery (44.7%) compared to patients receiving no surgery of the primary site (21.5%) (p < 0.001). Both local therapy and radical surgery were each independently associated with decreased CSM, with predicted 5-yr cumulative incidence of 45.4 and 43.3%, respectively, compared to 64.7% for patients receiving no surgery of the primary site (p < 0.001). Multivariate analyses demonstrated that primary site surgery was independently associated with better OS (local therapy, p = 0.037; radical surgery, p < 0.001) and decreased CSM (p = 0.003). Similar results were noted regardless of age, sex, T stage, N stage, and AJCC prognostic groups based on subgroup analysis. However, patients with M1 disease who underwent primary site surgery did not exhibit any survival benefit. CONCLUSION: Surgery for the primary tumor conferred a survival advantage in non-metastatic PUC patients.

Diagnostic Performance of Confocal Laser Endomicroscopy for the Detection of Bladder Cancer: Systematic Review and Meta-Analysis.

OBJECTIVE: To systematically evaluate the diagnostic efficacy of confocal laser endomicroscopy (CLE) in detection of bladder cancer. METHODS: A systematic literature search on CLE in diagnosing bladder cancer in PubMed, Embase, and the Cochrane Library databases was performed. A bivariate meta-regression model was used for meta-analysis to evaluate the pooled diagnostic value of CLE. RESULTS: A total of 5 eligible studies involving 302 lesions were available for this meta-analysis. In a per-lesion analysis, pooled sensitivity, specificity, positive likelihood ratio (PLR), negative likelihood ratio (NLR), diagnostic odds ratio (DOR), and summary receiver-operating curve (SROC) area under the curve (AUC) of CLE for malignant lesions were 0.90 (95% confidence interval [CI]: 0.85-0.94), 0.72 (95% CI: 0.59-0.82), 3.20 (95% CI: 2.14-4.79), 0.14 (95% CI: 0.09-0.21), 23.27 (95% CI: 11.71-46.25), and 0.91 (95% CI: 0.89-0.94), respectively. For low-grade urothelial carcinomas, pooled sensitivity, specificity, PLR, NLR, DOR, and AUC for CLE were 0.72 (95% CI: 0.57-0.84), 0.87 (95% CI: 0.77-0.93), 5.48 (95% CI: 3.12-9.62), 0.32 (95% CI: 0.20-0.50), 17.19 (95% CI: 8.01-36.89), and 0.85 (95% CI: 0.82-0.88), respectively. For high-grade urothelial carcinomas, pooled sensitivity, specificity, PLR, NLR, DOR, and AUC for CLE were 0.82 (95% CI: 0.62-0.92), 0.84 (95% CI: 0.73-0.91), 4.96 (95% CI: 2.58-9.54), 0.22 (95% CI: 0.09-0.52), 22.49 (95% CI: 5.33-94.85), and 0.89 (95% CI: 0.86-0.91), respectively. CONCLUSION: CLE is a promising endoscopy technique for real-time tumor grading of bladder cancer.

Resource reallocation patterns within Sagittaria trifolia inflorescences following differential pollination.

PREMISE OF THE STUDY: Understanding resource allocation to reproduction, a key factor in life history tradeoffs, has long intrigued plant ecologists. Despite the recognized importance of understanding the movement of resources among flowers following variable pollination, the patterns of resource reallocation to plant reproductive organs have not been thoroughly addressed. In this study, we aimed to empirically explore how resources redistribute within inflorescences in response to differential pollination intensities. METHODS: Using a common herb, Sagittaria trifolia, we conducted supplemental and controlled pollination for single, some, or all flowers in simple and complex inflorescences, and compared their resulting fruiting probabilities, seed production, and average seed masses. KEY RESULTS: Pollen supplementation of a single flower significantly increased its fruiting probability; however, the same manipulation of an inflorescence did not increase its overall reproduction. Single pollen-supplemented flowers had a higher percentage fruit set than inflorescences receiving supplemental pollination. In complex inflorescences, supplemental pollination had no effect on the reproductive success of flowers on the lateral or main branches. CONCLUSIONS: We provided evidence of resource reallocation from controlled to pollen-supplemented flowers in simple inflorescences; however, resources were unlikely to be reallocated between the main and lateral branches in the complex inflorescences, suggesting that flowering branches represent integrated physiological units in S. trifolia. The results also demonstrated that single-flower supplemental pollination would exaggerate pollen limitation and lead to a biased understanding of a plant's reproductive status.

Healing Wounds Efficiently with Biomimetic Soft Matter: Injectable Self-Healing Neutral Glycol Chitosan/Dibenzaldehyde-Terminated Poly(ethylene glycol) Hydrogel with Inherent Antibacterial Properties.

The high prevalence of acquiring skin wounds, along with the emergence of antibiotic-resistant strains that lead to infections, impose a threat to the physical, mental, and socioeconomic health of society. Among the wide array of wound dressings developed, hydrogels are regarded as a biomimetic soft matter of choice owing to their ability to provide a moist environment ideal for healing. Herein, neutral glycol chitosan (GC) was cross-linked via imine bonds with varying concentrations of dibenzaldehyde-terminated polyethylene glycol (DP) to give glycol chitosan/dibenzaldehyde-terminated polyethylene glycol hydrogels (GC/DP). These dynamic Schiff base linkages (absorption peak at 1638 cm-1) within the hydrogel structure endowed their ability to recover from damage as characterized by high-low strain exposure in continuous step strain rheology. Along with their good injectability and biodegradability, the hydrogels exhibited remarkable inhibition against E. coli, P. aeruginosa, and S. aureus. GC/DP hydrogels demonstrated high LC50 values in vivo using zebrafish embryos as a model system due to their relative biocompatibility and a remarkable 93.4 ± 0.88% wound contraction at 30-dpw against 49.1 ± 3.40% of the control. To the best of our knowledge, this is the first study that developed injectable glycol chitosan/dibenzaldehyde-terminated polyethylene glycol self-healing hydrogels for application in wound healing with intrinsic bacteriostatic properties against the three bacteria.

Molecular networking-assisted systematical profiling and the in vivo neuroprotective effect of ellagitannins from the Melastoma dodecandrum Lour.

BACKGROUND: Ellagitannins (ETs) are a major classification of natural tannins, with relatively large and complex structures. ETs from medicinal plants are focused increasingly due to urolithins, a kind of intestinal metabolite of ETs, which showed promising anti-Alzheimer's disease (AD) effects. Melastoma dodecandrum (MD), a widely used traditional Chinese medicine is rich in ETs, but their chemistry and potential neuroprotective effects have not been investigated. PURPOSE: This study aimed to identify the chemical composition of ETs in the crude extract of MD and to investigate their neuroprotective effects in vivo. METHODS: UPLC-QTOF-MS-based molecular networking (MN) and structural characterization were applied to targeted profiling of the MD-ETs. Animal behavior experiments, including the novel object recognition test (NOR), open field test (OFT), and Morris water maze test (MWM), were conducted to assess the memory improvement effects of MD-ETs in AD model mice. RESULTS: A total of 70 ETs, ranging from monomers to tetramers, were tracked and characterized in the MD extract using MN-guided targeted profiling, with 59 of them reported for the first time in this species. MD-ETs significantly improved memory impairment in AD mice, as indicated by decreased escape latency, increased number of crossings and target quadrant distance in MWM, increased rearing number in OFT, and increased preference index in NOR. CONCLUSION: This study systematically characterized the composition and structural features of ETs in MD using targeted LC-MS profiling, expanding the chemical information of ETs in MD. Furthermore, the results demonstrate that MD-ETs have significant effects on improving impaired memory in AD mice, suggesting their potential as alternative natural medicines for the treatment of neurodegenerative diseases.

[Literature study on the syndrome diagnosis and classification of coronary artery disease].

To investigate the application of diagnosis methods for coronary artery disease (CAD) and the status quo of its syndrome typing. The literature content analysis was used in this study. The frequency statistics was performed by comprehensively collecting CAD (thoracic obstruction) syndrome typing correlated literatures, designing content analysis tables, extracting information such as typing methods, typing elements, and syndrome typing, and so on. Results showed that blood stasis, yin deficiency, qi deficiency, yang deficiency, phlegm turbidity, and other syndrome elements extensively exist in literatures concerning CAD syndrome typing. Modem doctors prefer to use syndrome typing of qi, blood, jinye, and eight principles in syndrome typing of CAD more frequently. The asthenia in origin and asthenia in superficiality has been widely recognized as the basic syndrome of CAD.

Intravesical Recurrence After Radical Nephroureterectomy of Upper Urinary Tract Urothelial Carcinoma: A Large Population-Based Investigation of Clinicopathologic Characteristics and Survival Outcomes.

Background: Of patients with upper urinary tract urothelial carcinoma (UTUC), 22-47% developed bladder recurrence after radical nephroureterectomy. Furthermore, the effect of surgery for UTUC-bladder cancer (BC) has not been well validated. The aim of this study was to assess the impact of standard primary BC surgical strategy on survival of patients diagnosed with UTUC-BC. Patients and Methods: A total of 676 UTUC-BC patients and 197,753 primary BC patients diagnosed from 2004 to 2016, were identified based on the SEER database. The Kaplan-Meier method and the Fine and Gray competing risks analysis were performed to assess overall survival (OS) and cancer-specific mortality (CSM). Multivariate Cox regression model and competing risks regression model were used to identify independent risk factors. Propensity score matching (PSM) was also performed to adjust potential confounding factors. Results: The baseline characteristics and survival outcomes of the two BC patient cohorts are quite different. For UTUC-BC patients, no significant difference in OS (NMIBC: p = 0.88; MIBC: p = 0.98) or cumulative incidence of CSM (NMIBC: p = 0.12; MIBC: p = 0.96) were noted for various surgical procedures. Local tumor treatment and partial cystectomy for UTUC-NMIBC patients produced lower 1-year (6.1%) and 3-year CSM (16.2%). Radical cystectomy for UTUC-MIBC patients produced lower 1-year (11.8%) but higher 3-year CSM (62.7%). After PSM for covariates, UTUC-BC patients still had a worse prognosis after surgery compared with primary BC patients. Based on regression models, older age, advanced T stage, N positive disease, M positive disease, and shorter interval between UTUC and BC were identified as independent risk factors for UTUC-BC patients. Conclusion: Standard primary BC surgical strategy did not provide significant survival benefit for UTUC-BC patients. Compared with primary BC patients, UTUC-BC patients had a worse prognosis after surgery, suggesting that current primary BC surgical guidelines are not entirely appropriate for UTUC-BC patients. Our findings underscore the continued importance and need for better prognosis and improved guidelines for management of UTUC-BC patients.

Prognostic Value of an Immunohistochemical Signature in Patients With Bladder Cancer Undergoing Radical Cystectomy.

BACKGROUND: This study aimed to assess the prognostic value of various diagnostic immunohistochemical (IHC) markers and develop an IHC-based classifier to predict the disease-free survival (DFS) of patients with bladder cancer undergoing radical cystectomy. METHODS: IHC was performed on tumor specimens from 366 patients with transitional cell bladder cancer. The least absolute shrinkage and selection operator (LASSO) Cox regression model was used to develop a multi-marker classifier for predicting DFS of patients with bladder cancer. The Kaplan-Meier estimate was performed to assess DFS, and unadjusted and adjusted Cox regression models were used to identify independent risk factors to predict DFS of patients with bladder cancer. RESULTS: Based on the LASSO Cox regression model, nine prognostic markers were identified in the training cohort. Patients were stratified into low- and high-risk groups using the IHC-based classifier. In the training cohort, the 10-year DFS was significantly better in low-risk patients (71%) compared with high-risk patients (18%) (p < 0.001); in the validation cohort, the 10-year DFS was 86% for the low-risk group and 20% for the high-risk group (p < 0.001). Multivariable Cox regression analyses showed that the high-risk group based on the classifier was associated with poorer DFS adjusted by clinicopathological characteristics. Finally, a nomogram comprising the classifier and clinicopathological factors was developed for clinical application. CONCLUSION: The nine-IHC-based classifier is a reliable prognostic tool, which can eventually guide clinical decision making regarding treatment strategy and follow-up scheduling of bladder cancer.

Association of genetic variants in migraineurs with and without restless legs syndrome.

OBJECTIVE: Several single-nucleotide polymorphisms (SNPs) are associated with restless legs syndrome (RLS). This study investigated whether or not additional SNP variants increase the risk of RLS in migraineurs and in migraine with aura (MA) and migraine without aura (MoA) subgroups. METHODS: Migraineurs with and without RLS were genotyped using an Affymetrix array. We performed association analyses for the entire cohort and the MA and MoA subgroups, which were divided further into episodic migraine (EM) and chronic migraine (CM). Potential correlations between SNPs and clinical indices in migraineurs with RLS were examined by multivariate regression analysis. RESULTS: The rs77234324 and rs79004933 SNPs were found in migraineurs with (P = 2.57E-07) and without (P = 3.03E-07) RLS. The A allele frequency for rs77234324 (on LGR6) was 0.1321 in migraineurs with RLS and 0.0166 in those without RLS (odds ratio, 8.978). The T allele frequency for rs79004933 (in the intergenic region) was 0.1981 in migraineurs with RLS and 0.0446 in those without (odds ratio, 5.281). rs2858654, rs76770509, rs4243475 in UTRN, rs150762626, and rs2668375 were identified in migraine with and without RLS in the MoA subgroup (P = 7.56E-09, P = 2.30E-08, P = 1.19E-07, P = 6.86E-07, and P = 8.05E-07, respectively). There was a suggestion of an association between rs10510331 (P = 1.50E-06) and CM and EM in patients with MoA and RLS. Multivariate regression showed a significant relationship between rs79004933 and the Beck Depression Inventory score. INTERPRETATION: rs77234324 in LGR6 and rs79004933 in the intergenic region were associated with RLS in migraineurs. Five SNPs increased the risk of RLS in patients with MoA.

High Expression of CD39 is Associated with Poor Prognosis and Immune Infiltrates in Clear Cell Renal Cell Carcinoma.

INTRODUCTION: The cell-surface ectonucleotidase CD39 is a key molecule of the immunosuppressive adenosine pathway within the tumor microenvironment. However, the relationship between CD39 and clear cell renal cell carcinoma (ccRCC) is rarely reported and still remains unclear. METHODS: CD39 expression was first analyzed using the Oncomine and the Tumor IMmune Estimation Resource (TIMER) databases, and then examined in ccRCC patients (n=367) who had undergone radical nephrectomy using immunohistochemistry (IHC) and real-time quantitative PCR analysis (qPCR). The prognosis value of CD39 in ccRCC was evaluated by Cox proportional hazards analysis. Functional and gene set enrichment analysis (GSEA) was performed using transcriptomic data of ccRCC from TCGA. Correlation analysis between CD39 and tumor-infiltrating lymphocytes (TILs) was performed using the TISIDB database. The impact of CD39 on immune checkpoint therapy (ICT) was evaluated by two public cohorts. RESULTS: CD39 mRNA and protein expression was upregulated in tumor tissues from ccRCC patients and aberrant expression of CD39 was associated with advanced tumor stage and poor prognosis in ccRCC patients. EMT, IL-2/STAT5, inflammatory response, interferon gamma and KRAS hallmark gene sets were identified as CD39-related signaling pathway. The expression level of CD39 was significantly and positively correlated with high abundance of the regulatory TILs including NK cells, macrophages, Th cells and Treg cells. CD39 was correlated with expression of several immune checkpoints and higher CD39 expression was associated with better OS of ccRCC patients who received ICT. CONCLUSION: CD39 is a powerful prognostic marker of ccRCC patients. Increased tumor expression of CD39 mRNA is significantly correlated with infiltrating levels of TILs, and better efficacy of ICT to ccRCC. CD39 could be a novel therapeutic target for ccRCC.

JAK2 Mediates the Regulation of Pept1 Expression by Leptin in the Grass Carp (Ctenopharyngodon idella) Intestine.

Oligopeptide transporter 1 (Pept1) is located on the brush border membrane of the intestinal epithelium and plays an important role in dipeptide and tripeptide absorption from protein digestion. In this study, we cloned and characterized the cDNA sequence of Janus kinase 2 (JAK2) from Ctenopharyngodon idella. The expression patterns of JAK2 in various tissues and developmental stages were characterized by quantitative real-time PCR (qRT-PCR). The mRNA expression levels of JAK2 and Pept1 regulated by leptin in the intestine were also analyzed in vitro and in vivo. The cDNA sequence of JAK2 is 3378 bp in length, and the mRNA of JAK2 was broadly expressed in all tissues and embryonic stages of C. idella analyzed. In addition, we found that leptin regulated expression of JAK2 and Pept1 in the intestine; Pept1 expression was down-regulated by the JAK2 inhibitor AG490 in vivo and in vitro. Furthermore, luciferase experiments showed that overexpression of the JAK2 gene significantly upregulated the activity of the Pept1 5' regulatory sequence in C. idella. In conclusion, these results may help in elucidating the regulatory effect of the leptin-mediated JAK2 pathway on intestinal Pept1 expression in C. idella and the molecular mechanism of peptide transport by the intestinal transporter Pept1 in fishes.

[Cluster analysis on TCM syndromes in 319 coronary artery disease patients for establishment of syndrome diagnostic figure].

OBJECTIVE: To explore the diagnostic figures for TCM syndrome typing in coronary heart disease (CHD) patients. METHODS: A retrospective investigation was carried out in 319 CHD patients hospitalized from Jan. 2004 to Dec. 2004 in authors' hospital. Through cluster analysis, descriptive statistics and frequency normalization in combination of clinical observation, the diagnostic figures of TCM syndromes were obtained. RESULTS: The figures for qi deficiency syndrome were: primary symptoms: chest pain and stuffiness, secondary symptoms: tiredness, short breath, poor appetite, light colored tongue, deep and thready pulse; for qi deficiency with phlegm and blood stasis syndrome: primary symptoms: chest stuffiness and pain, secondary symptoms: tiredness, insomnia, palpitation, obesity, dark red tongue, string and slippery pulse; for turbid-phlegm blocking collateral syndrome: primary symptoms: chest stuffiness, secondary symptoms: cough, expectoration with much white sputum, tiredness, short breath and poor appetite, light colored tongue with white greasy coating, slippery pulse. CONCLUSION: Research on diagnostic criteria for TCM syndrome typing could be established upon clinical epidemiologic survey and statistic analysis in combining with specialists' suggestions to primarily set the referrence figures.

[Trial study on DENG Tie-tao's coronary heart disease capsules in improving patients' quality of life].

OBJECTIVE: To assess the efficacy of the Coronary Heart Disease (CHD) Capsules worked out by Prof. Deng --in improving quality of life of CHD patients of qi deficiency with phlegm and blood stasis syndrome. METHOD: According to the WHO's diagnosis criteria of CHD, a total of 93 stable angina patients were divided into 3 groups using the single blinded method. The groups were evenly distributed into CHD Capsule treated group (CHDC), isosorbide dinitrate control group (ID), and Compound Prescription Danshen Droplet Pills control group (CPDDP). Two courses of treatment lasting for 6 months were given. During the courses of treatment, the following parameters were observed: clinical symptoms of angina pectoris, ECG change, treadmill exercise test, 36 items in short form of health survey (SF-36) and Seattle Angina Questionnaire (SAQ) scale. RESULTS: After 6 months of treatment, all the three groups showed good curative effect in angina pectoris, ECG and treadmill exercise test, differences between them had no statistical significance. The CHDC group showed a better result in nitro-glycerine stopping or alleviation rate and in improving symptoms than the other groups (P < 0.05). The general health, vitality, role-emotional, mental health and reported health transition in the CHDC group were significantly better than those in the control groups (P < 0.05). The scores in physiological functioning role, physiological function and pain alleviation were not different among the three groups. CONCLUSION: Prof. DENG Tie-tao's CHDC is effective in treating CHD with qi deficiency, phlegm and blood stasis and also in improving the quality of life. CHDC is more suitable to be used in long-term treatment than isosorbide dinitrate. The SF-36 and SAQ can be used to appraise the curative effect of traditional Chinese medicine agents for CHD angina pectoris.

Oxazolone-induced murine model of ulcerative colitis.

OBJECTIVE: Animal models are useful for studying disease, but there is a shortage of suitable models of ulcerative colitis. The aim of the present study was to set up an oxazolone-induced murine colitis model and use it to research the pathogenesis of inflammatory bowel disease. METHODS: BALB/c mice were presensitized by painting the skin with 0.2 mL 3% oxazolone in 100% ethanol on days 0 and 1 followed by intrarectal administration of 0.15 mL 1% oxazolone in 50% ethanol on day 7. The disease activity index (DAI), histological changes of the colon, myeloperoxidase (MPO) activity and production of cytokines (TNF-alpha, IL-4, IFN-gamma) by the mucosa were evaluated. RESULTS: There were obvious changes in the DAI, histology and MPO activity, and the production of interleukin-4 was markedly increased compared with the concentrations of TNF-alpha and IFN-gamma, which remained normal, in the lesions. CONCLUSION: Oxazolone colitis is Th2-mediated and has similar histologic features and distribution of inflammation to ulcerative colitis (UC), which has important implications for the use of this model in the study of the pathogenesis and treatment of UC.

Nicastrin is critical for stability and trafficking but not association of other presenilin/gamma-secretase components.

gamma-Secretase, which is responsible for the intramembranous cleavage of Alzheimer beta-amyloid precursor protein and the signaling receptor Notch, is a multiprotein complex consisting of at least four components: presenilin (PS); nicastrin (Nct); APH-1 (anterior pharynx-defective-1); and presenilin enhancer-2 (PEN-2). Presenilin 1 (PS1) is known to be essential for the stability, interaction, and trafficking of the other PS1/gamma-secretase components. However, the precise functions of the other components remain elusive. Here, we investigated the functions of Nct within the PS1/gamma-secretase complex. We demonstrated that the loss of Nct expression in the embryonic fibroblast cells (Nct KO cells) results in dramatically decreased levels of APH-1, PEN-2, and PS1 fragments accompanied by a significant accumulation of full-length PS1. In the absence of Nct, PEN-2 and full-length PS1 are subjected to proteasome-mediated degradation, whereas the degradation of APH-1 is mediated by both proteasomal and lysosomal pathways. Unlike the case of wild type cells in which the gamma-secretase complex mainly locates in the trans-Golgi network, the majority of residual PEN-2, APH-1, and the uncleaved full-length PS1 in Nct KO cells reside in the endoplasmic reticulum, which remain associated with each other in the absence of Nct. Interestingly, significant amounts of full-length PS1 and PEN-2, but not APH-1, are detected on the plasma membrane in Nct KO cells, suggesting the Nct-independent cell surface delivery of the PEN-2.PS1. Finally, the diminished PEN-2 protein level in Nct-deficient cells can be partially restored by overexpression of exogenous PS1, APH-1, or PEN-2 individually or collectively, indicating a dispensable role for Nct in controlling PEN-2 level. Taken together, our study demonstrates a critical role of Nct in the stability and proper intracellular trafficking of other components of the PS1/ gamma-secretase complex but not in maintaining the association of PEN-2, APH-1, and full-length PS1.

An ER membrane protein, Sop4, facilitates ER export of the yeast plasma membrane [H+]ATPase, Pma1.

We have analyzed the mechanism by which Sop4, a novel ER membrane protein, regulates quality control and intracellular transport of Pma1-7, a mutant plasma membrane ATPase. At the restrictive temperature, newly synthesized Pma1-7 is targeted for vacuolar degradation instead of being correctly delivered to the cell surface. Loss of Sop4 at least partially corrects vacuolar mislocalization, allowing Pma1-7 routing to the plasma membrane. Ste2-3 is a mutant pheromone receptor which, like Pma1-7, is defective in targeting to the cell surface, resulting in a mating defect. sop4delta suppresses the mating defect of ste2-3 cells as well as the growth defect of pma1-7. Visualization of newly synthesized Pma1-7 in sop4delta cells by indirect immunofluorescence reveals delayed export from the ER. Similarly, ER export of wild-type Pma1 is delayed in the absence of Sop4 although intracellular transport of Gas1 and CPY is unaffected. These observations suggest a model in which a selective increase in ER residence time for Pma1-7 may allow it to achieve a more favorable conformation for subsequent delivery to the plasma membrane. In support of this model, newly synthesized Pma1-7 is also routed to the plasma membrane upon release from a general block of ER-to-Golgi transport in sec13-1 cells.

Presenilins and gamma-secretase inhibitors affect intracellular trafficking and cell surface localization of the gamma-secretase complex components.

The intramembranous cleavage of Alzheimer beta-amyloid precursor protein and the signaling receptor Notch is mediated by the presenilin (PS, PS1/PS2)-gamma-secretase complex, the components of which also include nicastrin, APH-1, and PEN-2. In addition to its essential role in gamma-secretase activity, we and others have reported that PS1 plays a role in intracellular trafficking of select membrane proteins including nicastrin. Here we examined the fate of PEN-2 in the absence of PS expression or gamma-secretase activity. We found that PEN-2 is retained in the endoplasmic reticulum and has a much shorter half-life in PS-deficient cells than in wild type cells, suggesting that PSs are required for maintaining the stability and proper subcellular trafficking of PEN-2. However, the function of PS in PEN-2 trafficking is distinct from its contribution to gamma-secretase activity because inhibition of gamma-secretase activity by gamma-secretase inhibitors did not affect the PEN-2 level or its egress from the endoplasmic reticulum. Instead, membrane-permeable gamma-secretase inhibitors, but not a membrane-impermeable derivative, markedly increased the cell surface levels of PS1 and PEN-2 without affecting that of nicastrin. In support of its role in PEN-2 trafficking, PS1 was also required for the gamma-secretase inhibitor-induced plasma membrane accumulation of PEN-2. We further showed that gamma-secretase inhibitors specifically accelerated the Golgi to the cell surface transport of PS1 and PEN-2. Taken together, we demonstrate an essential role for PSs in intracellular trafficking of the gamma-secretase components, and that selective gamma-secretase inhibitors differentially affect the trafficking of the gamma-secretase components, which may contribute to an inactivation of gamma-secretase.

PEN-2 and APH-1 coordinately regulate proteolytic processing of presenilin 1.

Presenilin (PS, PS1/PS2) complexes are known to be responsible for the intramembranous gamma-secretase cleavage of the beta-amyloid precursor protein and signaling receptor Notch. PS holoprotein undergoes endoproteolysis by an unknown enzymatic activity to generate NH(2)- and COOH-terminal fragments, a process that is required for the formation of the active and stable PS/-gamma-secretase complex. Biochemical and genetic studies have recently identified nicastrin, APH-1, and PEN-2 as essential cofactors that physically interact with PS1 and are necessary for the gamma-secretase activity. However, their precise function in regulating the PS complex and gamma-secretase activity remains unknown. Here, we demonstrate that endogenous PEN-2 preferentially interacts with PS1 holoprotein. Down-regulation of PEN-2 expression by small interfering RNA (siRNA) abolishes the endoproteolysis of PS1, whereas overexpression of PEN-2 promotes the production of PS1 fragments, indicating a critical role for PEN-2 in PS1 endoproteolysis. Interestingly, accumulation of full-length PS1 resulting from down-regulation of PEN-2 is alleviated by additional siRNA down-regulation of APH-1. Furthermore, overexpression of APH-1 facilitates PEN-2-mediated PS1 proteolysis, resulting in a significant increase in PS1 fragments. Our data reveal a direct role of PEN-2 in proteolytic cleavage of PS1 and a regulatory function of APH-1, in coordination with PEN-2, in the biogenesis of the PS1 complex.