Identification of potential miRNA biomarkers for traumatic osteonecrosis of femoral head.

Traumatic osteonecrosis of femoral head (TONFH) is a common orthopedic disease caused by physical injury in hip. However, the unclear pathogenesis mechanism of TONFH and lacking of simple noninvasive early diagnosis method cause the necessity of hip replacement for most patients with TONFH. In this study, we aimed to identify circulating microRNAs (miRNAs) by integrated bioinformatics analyses as potential biomarker of TONFH. mRNA expression profiles were downloaded from the Gene Expression Omnibus database. Then we combined two miRNA screen methods: Weighted gene co-expression network analysis and fold change based differentially expressed miRNAs analysis. As a result, we identified 14 key miRNAs as potential biomarkers for TONFH. Besides, 302 target genes of these miRNAs were obtained and the miRNA-mRNA interaction network was constructed. Furthermore, the results of Kyoto Encyclopedia of Gene and Genome pathway analysis, Gene Ontology function analysis, protein-protein interaction (PPI) network analysis and PPI network module analysis showed close correlation between these 14 key miRNAs and TONFH. Then we established receiver operating characteristic curves and identified 6-miRNA signature with highly diagnosis value including miR-93-5p (area under the curve [AUC] = 0.93), miR-1324 (AUC = 0.92), miR-4666a-3p (AUC = 0.92), miR-5011-3p (AUC = 0.92), and miR-320a (AUC = 0.89), miR-185-5p (AUC = 0.89). Finally, the results of quantitative real-time polymerase chain reaction confirmed the significantly higher expression of miR-93-5p and miR-320a in the serum of patients with ONFH. These circulating miRNAs could serve as candidate early diagnosis markers and potential treatment targets of TONFH.

Plasma levels of lysophosphatidic acid in ovarian cancer versus controls: a meta-analysis.

BACKGROUND: In this study, using a meta-analysis approach, we examined the correlation between serum levels of lysophosphastidic acid (LPA) and ovarian cancer (OC). METHODS: Relevant published studies were identified from multiple scientific literature databases by using a pre-determined electronic and manual search strategy. The search results were screened through a multi-step process to select high-quality case-control studies suitable for the present meta-analysis. Mean values and standardized mean differences (SMD) were calculated for plasma LPA levels. Two investigators independently extracted the data from the studies and performed data analysis using STATA software version 12.0 (Stata Corp, College Station, TX, USA). RESULTS: Nineteen case-control studies met our selection criteria and contained a total of 980 OC patients, 872 benign controls and 668 healthy controls. Our meta-analysis results revealed that the plasma levels of LPA in OC patients were significantly higher than benign controls (SMD = 2.36, 95% CI: 1.61-3.10, P < 0.001) and healthy controls (SMD = 2.32, 95% CI: 1.77-2.87, P < 0.001). Subgroup analysis by ethnicity showed that the plasma LPA levels in OC patients were significantly higher than the benign controls only in Asian populations (SMD = 2.52, 95% CI: 1.79-3.25, P < 0.001). However, a comparison between healthy controls and OC patients revealed that, in both Asians and Caucasians, the OC patients displayed significantly higher plasma LPA levels compared to healthy controls (all P < 0.05). CONCLUSION: Our meta-analysis showed strong evidence that a significantly higher plasma LPA levels are present in OC patients, compared to benign controls and healthy controls, and plasma LPA levels may be used as a biomarker or target of OC.

Interactions between the apolipoprotein a1/c3/a5 haplotypes and alcohol consumption on serum lipid levels.

BACKGROUND: The interactions between apolipoprotein (Apo) A1/C3/A5 haplotypes and alcohol consumption on serum lipid profiles have not been previously explored. The present study was undertaken to detect the polymorphisms of ApoA1 -75 bp G>A (rs1799837), ApoC3 3238C>G (rs5128), ApoA5 -1131T>C (rs662799), ApoA5 c.553G>T (rs2075291), and ApoA5 c.457G>A (rs3135507) and the interactions between their haplotypes and alcohol consumption on serum lipid levels. METHODS: Genotyping was performed in 1,030 unrelated subjects (516 nondrinkers and 514 drinkers) aged 15 to 89. The interactions between ApoA1/C3/A5 haplotypes and alcohol consumption on serum lipid levels were detected by factorial regression analysis after controlling for potential confounders. RESULTS: The frequencies of ApoC3 3238 CG/GG genotypes and ApoA1 -75 bp A allele in nondrinkers were higher in females than in males (p < 0.05). The frequencies of ApoC3 3238 CG/GG genotypes and G allele in drinkers were higher in females than in males (p < 0.05). The frequencies of ApoA1 -75 bp GA/AA genotypes and A allele in males were higher, and those of ApoC3 3238 CG/GG genotypes were lower in drinkers than in nondrinkers (p < 0.05 to 0.01). The frequency of ApoC3 3238 GG genotype in male drinkers was also higher in ≥25 g/d than in <25 g/d subgroups (p < 0.05). There were 11 haplotypes with a frequency >1% in our study population. The haplotypes of G-G-T-C-G (in the order of c.553G>T, c.457G>A, -1131T>C, 3238C>G, and -75 bp G>A), G-G-T-C-A, and G-G-C-G-G were shown consistent interactions with alcohol consumption to increase serum total cholesterol, high-density lipoprotein cholesterol (HDL-C), and ApoA1 levels (p < 0.05 to 0.001). The interactions between G-G-T-G-G (HDL-C and ApoA1), G-G-C-C-A (ApoA1), G-A-T-C-G (triglyceride), G-G-T-C-G (ApoA1/ApoB ratio), and G-G-C-G-G (ApoB) haplotypes and alcohol consumption on serum lipid levels were also detected (p < 0.05 to 0.001); the levels of these serum lipid parameters were significantly higher in drinkers than in nondrinkers. CONCLUSIONS: The differences in serum lipid parameters between drinkers and nondrinkers might partly result from different interactions between the ApoA1/C3/A5 haplotypes and alcohol consumption.

Effects of slope aspect and age class on diseases of Juglans regia in Wild Walnut Nature Conservation Area of Xinjiang, China.

To understand the disease situation of vulnerable plant Juglans regia, we investigated the proportions of four diseases of J. regia with different age classes in different slope aspects in the Wild Walnut Nature Conservation Area of Xinjiang. We analyzed the correlation of disease types and disease grades with the diameter at breast height (DBH) of J. regia and slope aspects. The results showed that the main diseases of J. regia in the reserve were walnut brown spot disease (95.8%), walnut deadwood disease (90.5%), walnut black spot disease (74.4%), and walnut rot disease (7.7%). J. regia was susceptible to walnut brown spot disease in all the four slope aspects, with walnut deadwood disease on shady and semi-shady slopes, walnut black spot disease on sunny and shady slopes, and walnut rot disease on semi-sunny and semi-shady slopes. The proportion of diseased plants with the four diseases decreased with the increases of disease grades (1-4 grades) in the four slope aspects. Across the four slope aspects, the middle-aged trees had the largest proportion of walnut deadwood disease, walnut black spot disease and walnut brown spot disease, followed by old trees and young trees. No diseased seedlings were found. Walnut rot disease only occurred in old trees. There was significant positive correlation between DBH of J. regia and the four diseases. Walnut black spot disease had significant negative correlation with slope aspect. There was correlation between different disease grades of walnut deadwood disease, walnut rot disease and walnut black spot disease with DBH and slope aspects.

Prognostic models for early and late tumor progression prediction in nasopharyngeal carcinoma: An analysis of 8292 endemic cases.

OBJECTIVES: The time for posttreatment tumor progression differs between nasopharyngeal carcinoma (NPC) patients. Herein, we established effective nomograms for predicting early tumor progression (ETP) and late tumor progression (LTP) in NPC patients. METHODS: We retrospectively enrolled 8292 NPC patients (training cohort: n = 6219; validation cohort: n = 2073). The ELP and LTP were defined as the time to tumor progression ≤24 and >24 months after treatment, respectively. RESULTS: The ETP and LTP accounted for 52.6 and 47.4% of the total patient cohort, respectively. Patients who developed ETP had markedly worse overall survival (OS) versus patients who suffered from LTP (5-year OS: 26.2% vs. 59.7%, p < 0.001). Further, we identified 10/6 predictive factors significantly associated with ETP/LTP via logistic regression analyses. These indicators were used separately to construct two predictive nomograms for ETP and LTP. In the training group, the ETP nomogram [Harrell Concordance Index (C-index) value: 0.711 vs. 0.618; p < 0.001] and LTP nomogram (C-index value: 0.701 vs. 0.612; p < 0.001) were significantly superior for risk stratification than the TNM staging. These results were supported in the validation group with a C-index value of 0.753 and 0.738 for the ETP and LTP nomograms, respectively. High-risk patients defined by ETP/LTP nomograms had shorter progression-free survival than low-risk patients (all p < 0.001). CONCLUSION: The established nomograms can help in ELP or LTP risk stratification for NPC patients. Our current results might also provide insights into individualized treatment decisions and designing surveillance strategies for NPC patients.

Ferroptosis patterns and tumor microenvironment infiltration characterization in esophageal squamous cell cancer.

Background: Esophageal Squamous Cell Cancer (ESCC) is an aggressive disease associated with a poor prognosis. As a newly defined form of regulated cell death, ferroptosis plays a crucial role in cancer development and treatment and might be a promising therapeutic target. However, the expression patterns of ferroptosis-related genes (FRGs) in ESCC remain to be systematically analyzed. Methods: First, we retrieved the transcriptional profile of ESCC from TCGA and GEO datasets (GSE47404, GSE23400, and GSE53625) and performed unsupervised clustering to identify different ferroptosis patterns. Then, we used the ssGSEA algorithm to estimate the immune cell infiltration of these patterns and explored the differences in immune cell abundance. Common genes among patterns were finally identified as signature genes of ferroptosis patterns. Results: Herein, we depicted the multi-omics landscape of FRGs through integrated bioinformatics analysis and identified three ESCC subtypes with distinct immune characteristics: clusters A-C. Cluster C was abundant in CD8+ T cells and other immune cell infiltration, while cluster A was immune-barren. By comparing the differently expressed genes between clusters of diverse datasets, we defined a gene signature for each cluster and successfully validated it in the TCGA-ESCC dataset. Conclusion: We provided a comprehensive insight into the expression pattern of ferroptosis genes and their interaction with immune cell infiltration. Additionally, we established a gene signature to define the ferroptosis patterns, which might be used to predict the response to immunotherapy.

CircRNA circUSP36 impairs the stability of NEDD4L mRNA through recruiting PTBP1 to enhance ULK1-mediated autophagic granulosa cell death.

BACKGROUND: Hypercholesterolemia is defined as a high risk factor for causing female infertility by changing the cholesterol level in granulosa cells to impair the microenvironment of oocyte development and maturation. High blood levels of oxidized low-density lipoprotein (ox-LDL) undergoes an increase of autophagic granulosa cell death. Unfortunately, this underlying molecular mechanism remains largely elusive. We aim to uncover the role of circ-ubiquitin specific peptidase 36 (USP36) in autophagic granulosa cell death. METHODS: Exposure of ox-LDL on the ovarian granulosa cell-like human granulosa (KGN) cells line was established for simulating the situation of hypercholesterolemia in vitro. Levels of circUSP36 and ULK1 were detected using real-time polymerase chain reaction (RT-PCR). Cell viability and apoptosis were assessed using (4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, flow cytometry and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining, respectively. Immunofluorescence staining of LC3 was performed to evaluate activity of autophagy. Western blot was employed to determine expression of apoptosis and autophagy-associated markers. RNA immunoprecipitation (RIP) and RNA pull-down assays were subjected to verify the circUSP36-PTBP1-NEDD4L regulatory axis. RESULTS: Treatment of ox-LDL induced aberrantly up-regulated circUSP36. Knockdown of circUSP36 alleviated cell apoptosis and excessive autophagy of granulosa cells triggered by ox-LDL. Mechanistically, reinforced expression of circUSP36 guided and facilitated PTBP1 binding to the coding region (CDS) of NEDD4L, resulting in NEDD4L mRNA decay. ULK1 was regulated by the circUSP36-PTBP1-NEDD4L axis in granulosa cells, thereby contributing to autophagic granulosa cell death. CONCLUSIONS: In summary, ox-LDL fostered autophagic granulosa cell death through circUSP36-mediated NEDD4L mRNA decay, thus elevating ULK1 expression.

Expression of macrophage migration inhibitory factor and CD74 in cervical squamous cell carcinoma.

OBJECTIVE: Macrophage migration inhibitory factor (MIF) and CD74 emerge as important players in pathogenesis and angiogenesis of several types of malignant tumors. The purpose of this study was to evaluate the expression of MIF and CD74 in cervical squamous cell carcinoma and explore the potential roles they play in cervical tumor angiogenesis. METHODS: Macrophage migration inhibitory factor and CD74 expression was assessed by immunohistochemistry in 209 cases with various degrees of cervical epithelial lesions, including 40 normal cervical epithelia, 43 mild cervical intraepithelial neoplasia 1 (CIN 1), 41 moderate-severe cervical intraepithelial neoplasia (CIN 2 to 3), and 85 cervical squamous cell carcinomas (SCCs). CD34 staining was used for counting microvessel density. Semiquantitative reverse transcription polymerase chain reaction and Western blot were used to detect messenger RNA and protein levels of MIF and CD74 in normal and malignant cervical tissues and cervical cancer cell lines SiHa and C-33A. The concentration of vascular endothelial growth factor (VEGF) in the conditioned media of cervical cancer cells was analyzed by enzyme-linked immunosorbent assay. RESULTS: Immunohistochemical analysis showed that MIF and CD74 expression was significantly higher in CIN than in the normal samples and higher in SCC than in CIN. The overexpression of MIF was correlated with deep stromal infiltration but not with the other clinicopathologic features of SCC. Correlation analyses revealed that MIF was positively related to CD74, and both protein levels were associated with microvessel density. Exogenous MIF induced VEGF secretion in SiHa and C-33A cells in a dose-dependent manner, which can be inhibited by MIF-specific inhibitor (ISO-1) or anti-CD74 antibody. CONCLUSION: Overexpression of MIF and CD74 in SCC and its precancerous lesions and the up-regulation of VEGF secretion in cervical cancer cells indicate that MIF and CD74 may play critical roles in the pathogenesis and angiogenesis of cervical cancer.

Apolipoprotein A1/C3/A5 haplotypes and serum lipid levels.

BACKGROUND: The association of single nucleotide polymorphisms (SNPs) in the apolipoprotein (Apo) A1/C3/A4/A5 gene cluster and serum lipid profiles is inconsistent. The present study was undertaken to detect the association between the ApoA1/C3/A5 gene polymorphisms and their haplotypes with serum lipid levels in the general Chinese population. METHODS: A total of 1030 unrelated subjects (492 males and 538 females) aged 15-89 were randomly selected from our previous stratified randomized cluster samples. Genotyping of the ApoA1 -75 bp G>A, ApoC3 3238C>G, ApoA5 -1131T>C, ApoA5 c.553G>T and ApoA5 c.457G>A was performed by polymerse chain reaction and restriction fragment length polymorphism combined with gel electrophoresis, and then confirmed by direct sequencing. Pair-wise linkage disequilibria and haplotype analysis among the five SNPs were estimated. RESULTS: The levels of high-density lipoprotein cholesterol (HDL-C) and ApoA1 were lower in males than in femailes (P < 0.05 for each). The allelic and genotypic frequencies of the SNPs were no significant difference between males and females except ApoC3 3238C>G. There were 11 haplotypes with a frequency >1% identified in the cluster in our population. At the global level, the haplotypes comprised of all five SNPs were significantly associated with all seven lipid traits. In particular, haplotype G-G-C-C-A (6%; in the order of ApoA5 c.553G>T, ApoA5 c.457G>A, ApoA5 -1131T>C, ApoC3 3238C>G, and ApoA1 -75bp G>A) and G-A-T-C-G (4%) showed consistent association with total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), ApoA1, ApoB, and the ApoA1/ApoB ratio. In addition, carriers of haplotype G-G-T-C-G (26%) had increased serum concentration of HDL-C and ApoA1, whereas carriers of G-G-C-G-G (15%) had high concentrations of TC, triglyceride (TG) and ApoB. We also found that haplotypes with five SNPs explain much more serum lipid variation than any single SNP alone, especially for TG (4.4% for haplotype vs. 2.4% for -1131T>C max based on R-square) and HDL-C (5.1% for haplotype vs. 0.9% for c.553G>T based on R-square). Serum lipid parameters were also correlated with genotypes and several environment factors. CONCLUSIONS: Several common SNPs and their haplotypes in the ApoA1/C3/A5 gene cluster are closely associated with modifications of serum lipid parameters in the general Chinese population.

Protective role of salidroside against aging in a mouse model induced by D-galactose.

OBJECTIVE: To investigate the protective effects of putative AGEs (advanced glycation endproducts) inhibitor salidroside against aging in an accelerated mouse aging model induced by D-galactose. METHODS: A group of 5-month-old C57BL/6J mice were treated daily with D-galactose, D-galactose combined with salidroside, salidroside alone, and control buffer for 8 weeks. At the end of the treatment, serum AGEs levels, neurological activities, expression of glial fibrillary acidic protein (GFAP) and neurotrophin-3 (NT-3) in the cerebral cortex, as well as lymphocyte proliferation and IL-2 production were determined. RESULTS: D-galactose induced mouse aging model was developed as described before. As expected, salidroside blocked D-galactose induced increase of serum AGEs levels. It also reversed D-galactose induced aging effects in neural and immune system, as evidenced by improving motor activity, increasing memory latency time, and enhancing lymphocyte mitogenesis and interleukin-2 (IL-2) production. Furthermore, elevated expression of GFAP and NT-3 in the aged model mice was also reduced upon salidroside treatment. CONCLUSION: Salidroside inhibits AGEs formation in vivo, which at least partially contributes to its anti-aging effect in D-galactose induced aging model.

Deep Insights into Gut Microbiota in Four Carnivorous Coral Reef Fishes from the South China Sea.

Investigations of gut microbial diversity among fish to provide baseline data for wild marine fish, especially the carnivorous coral reef fishes of the South China Sea, are lacking. The present study investigated the gut microbiota of four carnivorous coral reef fishes, including Oxycheilinus unifasciatus, Cephalopholis urodeta, Lutjanus kasmira, and Gnathodentex aurolineatus, from the South China Sea for the first time using high-throughput Illumina sequencing. Proteobacteria, Firmicutes, and Bacteroidetes constituted 98% of the gut microbiota of the four fishes, and 20 of the gut microbial genera recovered in this study represent new reports from marine fishes. Comparative analysis indicated that the four fishes shared a similar microbial community, suggesting that diet type (carnivorous) might play a more important role in shaping the gut microbiota of coral reef fishes than the species of fish. Furthermore, the genera Psychrobacter, Escherichia-Shigella, and Vibrio constituted the core microbial community of the four fishes, accounting for 61-91% of the total sequences in each fish. The lack of the genus Epulopiscium in the four fishes was in sharp contrast to what has been found in coral reef fishes from the Red Sea, in which Epulopiscium was shown to be the most dominant gut microbial genus in seven herbivorous coral reef fishes. In addition, while unique gut microbial genera accounted for a small proportion (8-13%) of the total sequences, many such genera were distributed in each coral reef fish species, including several genera (Endozoicomonas, Clostridium, and Staphylococcus) that are frequently found in marine fishes and 11 new reports of gut microbes in marine fishes. The present study expands our knowledge of the diversity and specificity of gut microbes associated with coral reef fishes.

Development and validation of a prognostic nomogram for the pre-treatment prediction of early metachronous metastasis in endemic nasopharyngeal carcinoma: a big-data intelligence platform-based analysis.

BACKGROUND: Early failure of cancer treatment generally indicates a poor prognosis. Here, we aim to develop and validate a pre-treatment nomogram to predict early metachronous metastasis (EMM) in nasopharyngeal carcinoma (NPC). METHODS: From 2009 to 2015, a total of 9461 patients with NPC (training cohort: n = 7096; validation cohort: n = 2365) were identified from an institutional big-data research platform. EMM was defined as time to metastasis within 2 years after treatment. Early metachronous distant metastasis-free survival (EM-DMFS) was the primary endpoint. A nomogram was established with the significant prognostic factors for EM-DMFS determined by multivariate Cox regression analyses in the training cohort. The Harrell Concordance Index (C-index), area under the receiver operator characteristic curve (AUC), and calibration curves were applied to evaluate this model. RESULTS: EMM account for 73.5% of the total metachronous metastasis rate and is associated with poor long-term survival in NPC. The final nomogram, which included six clinical variables, achieved satisfactory discriminative performance and significantly outperformed the traditional tumor-node-metastasis (TNM) classification for predicting EM-DMFS: C-index: 0.721 versus 0.638, p < 0.001; AUC: 0.730 versus 0.644, p < 0.001. The calibration curves showed excellent agreement between the predicted and actual EM-DMFS. The nomogram can stratify patients into three risk groups with distinct EM-DMFS (2-year DMFS: 96.8% versus 90.1% versus 80.3%, p < 0.001). A validation cohort supported the results. The three identified risk groups are correlated with the efficacy of different treatment regimens. CONCLUSION: Our established nomogram can reliably predict EMM in patients with NPC and might aid in formulating risk-adapted treatment decisions and personalized patient follow-up strategies.

Development of a Nomogram Model for Treatment of Nonmetastatic Nasopharyngeal Carcinoma.

Importance: Because of tumor heterogeneity, overall survival (OS) differs significantly among individuals with nasopharyngeal carcinoma (NPC), even among those with the same clinical stage. Relying solely on TNM staging to guide treatment remains imperfect. Objectives: To establish a comprehensive nomogram to estimate individualized OS and to explore stratified treatment regimens for risk subgroups in nonmetastatic NPC. Design, Setting, and Participants: This cohort study included 8093 patients diagnosed with NPC at a single center in China from April 2009 to December 2015. The sample was split into a training cohort (5398 participants [66.7%]) and validation cohort (2695 [33.3%]). Data were analyzed in May 2020. Exposures: Age, T stage, N stage, Epstein-Barr virus (EBV) DNA level, serum lactate dehydrogenase (LDH) levels, and albumin (ALB) levels. Main Outcomes and Measures: The primary end point was OS. The nomogram for estimating OS was generated based on multivariate Cox proportional hazards regression. The performance of the nomogram was quantified using Harrell concordance index (C index), the area under the curve (AUC) of the receiver operating characteristic curve, and a calibration curve. OS rates were established using the Kaplan-Meier method, and intersubgroup differences were examined by the log-rank test. Results: Among the 8093 participants, 5688 (70.3%) were men, and the median age at diagnosis was 45 years (range, 7-85 years). Six variables (age, T stage, N stage, EBV DNA levels, LDH levels, and ALB levels) were identified through multivariate Cox regression and incorporated into a nomogram to estimate OS. The resulting nomogram showed excellent discriminative ability and significantly outperformed the eighth edition of the American Joint Committee on Cancer/Union for International Cancer Control TNM staging system for estimating OS (C index, 0.716 [95% CI, 0.698-0.734] vs 0.643 [95% CI, 0.624-0.661]; P < .001; AUC, 0.717 [95% CI, 0.698-0.737] vs 0.643 [95% CI, 0.623-0.662]; P < .001), and the calibration curves showed satisfactory agreement between the actual and nomogram-estimated OS rates. The validation cohort confirmed the results. Patients were stratified into 4 risk groups based on the 25th, 50th, and 75th percentile score values estimated from the nomogram. The 4 nomogram-defined risk groups demonstrated significantly different intergroup OS (3-year OS rates: risk group 1, 1328 of 1345 [98.7%]; risk group 2, 1289 of 1341 [96.1%]; risk group 3, 1222 of 1321 [92.5%]; risk group 4, 1173 of 1391 [84.3%]; P < .001). These risk groups were associated with the efficacy of different treatment regimens. For example, for risk group 4, induction chemotherapy with concurrent chemoradiotherapy was associated with a significantly better OS than concurrent chemoradiotherapy (log-rank P = .008) and intensity-modulated radiotherapy alone (log-rank P < .001). Conclusions and Relevance: In this study, the proposed nomogram model enabled individualized prognostication of OS and could help to guide risk-adapted treatment for patients with nonmetastatic NPC.

Risk Assessment of Secondary Primary Malignancies in Nasopharyngeal Carcinoma: A Big-Data Intelligence Platform-Based Analysis of 6,377 Long-term Survivors from an Endemic Area Treated with Intensity-Modulated Radiation Therapy during 2003-2013.

PURPOSE: The incidence, risk factors and survival impact of secondary primary malignancies (SPMs) among survivors of nasopharyngeal carcinoma (NPC) treated with definitive intensity-modulated radiation therapy (IMRT) with or without chemotherapy are poorly characterized. METHODS AND MATERIALS: Consecutive patients (n=6,377) from the big-data intelligence platform at Sun Yat-sen University Cancer Center, China (in a high-incidence area) with newly diagnosed non-metastatic pathologically proven non-keratinizing undifferentiated NPC treated with IMRT±chemotherapy between January 2003 and June 2013 were retrospectively analyzed. Cumulative incidence of SPMs was calculated using the Kaplan-Meier method. Cox proportional hazards model was used to identify potential risk factors for SPMs and assess whether SPMs affect overall survival. RESULTS: Of the 6,377 patients, 189 (3.0%) suffered SPMs (median follow-up, 62 months). One-, 2-, 3-, 4-, and 5-cumulative risks of SPMs were 0.4%, 0.9%, 1.6%, 2.2%, and 2.6%, respectively. Latency from start of IMRT to SPMs diagnosis was 37 months (range, 6 to 102 months). In patients with SPMs, 14.3% suffered SPMs within 1 year post-IMRT: 1-3 years, 38.1%; 3-5 years, 33.9%; and >5 years, 13.7%. Lung cancer was the most common SPM (50/6,377, 0.78%). Multivariate analysis demonstrated sex (male, 64% increase), age (≥50 years, 68% increase), and smoking history (41% increase) were significant risk factors for SPMs, and SPMs were associated with poorer overall survival. CONCLUSION: This large cohort study confirms SPMs a dreadful complication for long-term survivors of NPC treated with IMRT. SPMs negatively impact overall survival in NPC. Close follow-up is recommended for older male survivors with a smoking history.

The identification of a small molecule inhibitor that specifically reduces T cell-mediated adaptive but not LPS-mediated innate immunity by T cell membrane-monocyte contact bioassay.

Proinflammatory cytokines such as TNFalpha and IL-1beta are produced in lesional skin of chronic plaque psoriasis patients, and at other sites of chronic inflammation such as arthritic joints. They play vital roles in maintaining inflammation. It has recently been suggested that activated T cell contact-mediated monocyte activation, leading to the production of proinflammatory cytokines, contributes to the pathogenesis of psoriasis and other chronic inflammatory diseases such as psoriatic arthritis and rheumatoid arthritis. Using a T cell membrane-monocyte contact bioassay, we have identified small molecule antagonists that differentially block anti-CD3/anti-CD28 activated T cell-mediated, but not LPS-stimulated, TNFalpha production from monocytes. We selected several kinase inhibitors from the Berlex/Schering kinase library and tested the effect of these compounds in blocking TNFalpha production in the T cell membrane-monocyte contact bioassay. We have demonstrated that one compound BLX-1, from a p38 MAP kinase inhibitor project, inhibited T cell-mediated TNFalpha production from monocytes by about 80%, without any effect on TNFalpha production from LPS-stimulated monocytes. Other BLX-1 analogs showed 32-83% inhibition of TNFalpha production with LPS stimulation as compared to almost 100% inhibition of T cell-mediated TNFalpha production. In contrast, PKC inhibitors BLX-5, Go6983, and Ro-31-8220, inhibited TNFalpha production from both activated T cell membrane- and LPS-stimulated monocytes to the same extent (in the range of 50-100% inhibition). Therefore, the activated T cell membrane-monocyte contact bioassay can be used to screen small molecule antagonists that specifically target adaptive but not LPS-mediated innate immunity. Small molecule TNFalpha inhibitors interfering specifically with activated T cell contact-mediated TNFalpha production from monocytes, but not with LPS-mediated TNFalpha production of myeloid cells, are predicted to have an improved side-effect profile and thus may provide more favorable therapeutics for the treatment of T cell-mediated inflammatory diseases.

CCR4 and CCR10 ligands play additive roles in mouse contact hypersensitivity.

Psoriasis, atopic dermatitis and allergic contact dermatitis are T-cell-mediated inflammatory skin diseases; chemokine receptors (CCR) 4 and 10 play an important role in the ligand-mediated recruitment of T cells into the skin in mice and humans, specifically with regards to tethering, firm adhesion and subsequent extravasation to the sight of injury. We utilized established murine models of dinitrofluorobenzene-, trimellitic acid anhydride- or oxazalone-induced contact hypersensitivity, to reflect the various Th-polarizations of different skin diseases, and investigated the functional effect of antibody blocking of single CCR ligands or combination therapy to block all CCR4 and CCR10 ligands. Our results indicate a greater reduction in inflammatory response--measured by oedema formation, myeloid cell and neutrophil infiltration and activity and CD3+ cell infiltration at the site of injury--with combination antibody therapy to CCR4 and CCR10 ligands versus controls, in nearly every tested condition. We conclude that blocking CCR4 and CCR10 simultaneously, or their ligands, should be beneficial in the treatment of T-cell-mediated skin diseases.

Targeting leukocyte recruitment in the treatment of psoriasis.

Tissue-selective recruitment of immunocytes to cutaneous tissues, a complex multistep cascade mediated by a large variety of cytokines, chemokines, and adhesion molecules, is thought to be a pivotal process in the pathogenesis of psoriasis. Following this notion, specifically targeting leukocyte trafficking remains an attractive approach for the treatment of psoriasis. It is increasingly recognized that during the pathogenesis of psoriasis not only effector T cells play important roles, but also multiple interactions between T cells, dendritic cells, macrophages, mast cells, endothelial cells, and keratinocytes are crucial for the full-fledged development of psoriasis. Meanwhile, the first biologics specifically inhibiting key molecules involved in cutaneous leukocyte recruitment have been approved for the treatment of psoriasis. It is, however, challenging to predict that molecules in this complex process with many redundant and/or functionally overlapping players will suffice as therapeutic targets. In this review, we will discuss the molecules and mechanisms involved in trafficking of different types of leukocytes and elucidate modes of action as well as therapeutic strategies of existing drugs and drug candidates.

Promising clinical outcomes of sequential and "Sandwich" chemotherapy and extended involved-field intensity-modulated radiotherapy in patients with stage IE /IIE extranodal natural killer/T-cell lymphoma.

BACKGROUND: The optimal treatment for the rare subtype of non-Hodgkin lymphoma, extranodal natural killer/T-cell lymphoma (ENKTL), nasal-type, has not been clearly defined. The purpose of the study was to investigate the efficacy of sequential and "Sandwich" chemotherapy and extended involved-field intensity-modulated radiotherapy (IMRT) in patients with stage IE /IIE extranodal ENKTL, nasal-type. METHODS: One hundred and fifty-five patients with stage IE /IIE nasal-type ENKTL were enrolled in the study, including 99 patients treated with sequential chemotherapy and extended involved-field IMRT (SCRT) and 56 patients with "Sandwich" chemotherapy and extended involved-field IMRT and chemotherapy (SCRCT). All patients were treated with extended involved-field IMRT with median dose of 54.6 Gy to the primary tumor and positive lymph nodes. Ninety-four patients had Ann Arbor stage IE disease, and 61 patients had stage IIE disease. RESULTS: The 5-year rates of loco-regional recurrence (LRR), progression-free survival (PFS), and overall survival (OS) were 17.0%, 78.5%, and 84.7%, respectively. Univariate analysis revealed that EBV DNA copy after treatment (normal vs elevated level) was significant prognostic factor for LRR, PFS, and OS (P < 0.001); therapeutic method (SCRT vs SCRCT) was significant prognostic factor for PFS (71.0% vs 91.8%, P = 0.011), but there was no significant effect on 5-year LRR and OS (22.2% vs 8.2%, P = 0.051 for LRR; 80.9% vs 91.8%, P = 0.199 for OS). CONCLUSIONS: Compared with SCRT, SCRCT was significantly associated with higher PFS rates and showed a trend toward improved loco-regional control. EBV DNA copy after treatment is a good index for recurrence and prognosis for stage IE /IIE ENKTL patients.

Proposal of a Pretreatment Nomogram for Predicting Local Recurrence after Intensity-Modulated Radiation Therapy in T4 Nasopharyngeal Carcinoma: A Retrospective Review of 415 Chinese Patients.

PURPOSE: Local relapse-free survival (LRFS) differs widely among patients with T4 category nasopharyngeal carcinoma (NPC). We aimed to build a nomogram incorporating clinicopathological information to predict LRFS in T4 NPC after definitive intensity-modulated radiation therapy (IMRT). MATERIALS AND METHODS: Retrospective study of 415 Chinese patients with non-metastatic T4 NPC treated with definitive IMRT with or without chemotherapy at our cancer center between October 2009 and September 2013. The nomogram for LRFS at 3 and 5 years was generated based on multivariate Cox proportional hazards regression, and validated using bootstrap resampling, assessing discriminative performance using the concordance index (C-index) and determining calibration ability via calibration curves. RESULTS: Five-year LRFS was 88.8%. We identified and incorporated four independent prognostic factors for LRFS: ethmoid sinus invasion, primary gross tumor volume, age, and pretreatment body mass index. The C-index of the nomogram for local recurrence was 0.732 (95% confidence interval, 0.726 to 0.738), indicating excellent predictive accuracy. The calibration curve revealed excellent agreement between nomogram-predicted and observed LRFS probabilities. Risk subgroups based on total point score cutoff values enabled effective discrimination of LRFS. CONCLUSION: This pretreatment nomogram enables clinicians to accurately predict LRFS in T4 NPC after definitive IMRT, and could help to facilitate personalized patient counselling and treatment strategies.

Mouse splenic B lymphocyte activation using different activation stimuli induces in vitro splicing of tumor necrosis factor-alpha nuclear pre-mRNA.

The pleiotropic functions of tumor necrosis factor-alpha (TNFalpha) have brought considerable attention in the past decade to its physiological and pathological roles in inflammatory and autoimmune diseases. However, little is known about how the production of TNFalpha is regulated at the transcriptional and translational levels in immune cells such as T and B lymphocytes. Our previous study demonstrated that unspliced "pre-mRNA" of TNFalpha is present in resting T cells. Initiation of splicing of TNFalpha pre-mRNA to mature mRNA requires T cell activation, which is unique and necessary for TNFalpha production when compared to its production in mononuclear phagocytes, including different lineages of macrophages (Mvarphi) and dendritic cells (DC). In this study, we further demonstrate that resting mouse B cells also contain pre-existing TNFalpha mRNA. The physiological process of B cell activation induced by (1) either the cross-linking of the B cell receptor (BCR) or CD40, (2) treatment with LPS, or PMA plus ionomycin, induces TNFalpha mRNA splicing in vitro. The kinetic response of TNFalpha splicing in B cells is much slower when compared to that in activated T cells. Studies using well-known kinase inhibitors demonstrated that MAP kinase kinase (MEK) and protein kinase C (PKC) are required for TNFalpha splicing upon stimulation through the BCR. These studies demonstrate that the production of TNFalpha in activated B cells is regulated differently than in activated T cells, and these differences may allow for the selective inhibition of TNFalpha in various autoimmune diseases depending on the mechanism of action of the selected anti-TNFalpha therapy.