Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field

Joint analysis of three genome-wide association studies of esophageal squamous cell carcinoma in Chinese populations

We conducted a joint (pooled) analysis of three genome-wide association studies (GWAS) 1-3 of esophageal squamous cell carcinoma (ESCC) in ethnic Chinese (5,337 ESCC cases and 5,787 controls) with 9,654 ESCC cases and 10,058 controls for follow-up. In a logistic regression model adjusted for age, sex, study, and two eigenvectors, two new loci achieved genome-wide significance, marked by rs7447927 at 5q31.2 (per-allele odds ratio (OR) = 0.85, 95% CI 0.82-0.88; P=7.72x10−20) and rs1642764 at 17p13.1 (per-allele OR= 0.88, 95% CI 0.85-0.91; P=3.10x10−13). rs7447927 is a synonymous single nucleotide polymorphism (SNP) in TMEM173 and rs1642764 is an intronic SNP in ATP1B2, near TP53. Furthermore, a locus in the HLA class II region at 6p21.32 (rs35597309) achieved genome-wide significance in the two populations at highest risk for ESSC (OR=1.33, 95% CI 1.22-1.46; P=1.99x10−10). Our joint analysis identified new ESCC susceptibility loci overall as well as a new locus unique to the ESCC high risk Taihang Mountain region

Long Noncoding RNA SOX2-OT: Regulations, Functions, and Roles on Mental Illnesses, Cancers, and Diabetic Complications

SRY-box transcription factor 2 (SOX2) overlapping transcript (SOX2-OT) is an evolutionarily conserved long noncoding RNA. Its intronic region contains the SOX2 gene, the major regulator of the pluripotency of embryonic stem cells. The human SOX2-OT gene comprises multiple exons and has multiple transcription start sites and generates hundreds of transcripts. Transcription factors (IRF4, AR, and SOX3), transcriptional inhibitors (NSPc1, MTA3, and YY1), and miRNAs (miR-211 and miR-375) have been demonstrated to control certain SOX2-OT transcript level at the transcriptional or posttranscriptional levels. Accumulated evidence indicates its crucial roles in the regulation of the SOX2 gene, miRNAs, and transcriptional process. Restricted expression of SOX2-OT transcripts in the brain results in the association between SOX2-OT single nucleotide polymorphisms and mental illnesses such as schizophrenia and anorexia nervosa. SOX2-OT is notably elevated in tumor tissues, and a high level of SOX2-OT is well correlated with poor clinical outcomes in cancer patients, leading to the establishment of its role as an oncogene and a prognostic or diagnostic biomarker for cancers. The emerging evidence supports that SOX2-OT mediates diabetic complications. In summary, SOX2-OT has diversified functions and could be a therapeutic target for various diseases

Preoperative serum immunoglobulin G and A antibodies to Porphyromonas gingivalis are potential serum biomarkers for the diagnosis and prognosis of esophageal squamous cell carcinoma

Abstract Background The key-stone-pathogen, Porphyromonas gingivalis associates not only with periodontal diseases but with a variety of other chronic diseases such as cancer. We previously reported an association between the presence of Porphyromonas gingivalis in esophageal squamous cell carcinoma (ESCC) and its progression. We now report the diagnostic and prognostic potential of serum immunoglobulin G and A antibodies (IgG/A) against Porphyromonas gingivalis for ESCC. Methods An enzyme-linked immunosorbent assay (ELISA) was used to determine the serum levels of Porphyromonas gingivalis IgG and IgA in 96 cases with ESCC, 50 cases with esophagitis and 80 healthy controls. Results The median serum levels of IgG and IgA for P. gingivalis were significantly higher in ESCC patients than non-ESCC controls. P. gingivalis IgG and IgA in serum demonstrated sensitivities/specificities of 29.17%/96.90% and 52.10%/70.81%, respectively, and combination of IgG and IgA produced a sensitivity/specificity of 68.75%/68.46%. The diagnostic performance of serum P. gingivalis IgA for early ESCC was superior to that of IgG (54.54% vs. 20.45%). Furthermore, high serum levels of P. gingivalis IgG or IgA were associated with worse prognosis of ESCC patients, in particular for patients with stage 0-IIor negative lymphnode metastasis, and ESCC patients with high levels of both IgG and IgA had the worst prognosis. Multivariate analysis revealed that lymph node status, IgG and IgA were independent prognostic factors. Conclusions The IgG and IgA for P. gingivalis are potential serum biomarkers for ESCC and combination of IgG and IgA improves the diagnostic and prognostic performance. Furthermore, serum P. gingivalis IgG and IgA can detect early stage ESCC

N-linked glycoproteomic profiling in esophageal squamous cell carcinoma

BACKGROUND: Mass spectrometry-based proteomics and glycomics reveal post-translational modifications providing significant biological insights beyond the scope of genomic sequencing. AIM: To characterize the N-linked glycoproteomic profile in esophageal squamous cell carcinoma (ESCC) via two complementary approaches. METHODS: Using tandem multilectin affinity chromatography for enrichment of N-linked glycoproteins, we performed N-linked glycoproteomic profiling in ESCC tissues by two-dimensional gel electrophoresis (2-DE)-based and isobaric tags for relative and absolute quantification (iTRAQ) labeling-based mass spectrometry quantitation in parallel, followed by validation of candidate glycoprotein biomarkers by Western blot. RESULTS: 2-DE-based and iTRAQ labeling-based quantitation identified 24 and 402 differentially expressed N-linked glycoproteins, respectively, with 15 in common, demonstrating the outperformance of iTRAQ labeling-based quantitation over 2-DE and complementarity of these two approaches. Proteomaps showed the distinct compositions of functional categories between proteins and glycoproteins with differential expression associated with ESCC. Western blot analysis validated the up-regulation of total procathepsin D and high-mannose procathepsin D, and the down-regulation of total haptoglobin, high-mannose clusterin, and GlcNAc/sialic acid-containing fraction of 14-3-3ζ in ESCC tissues. The serum levels of glycosylated fractions of clusterin, proline-arginine-rich end leucine-rich repeat protein, and haptoglobin in patients with ESCC were remarkably higher than those in healthy controls. CONCLUSION: Our study provides insights into the aberrant N-linked glycoproteome associated with ESCC, which will be a valuable resource for future investigations

Cascarinoids A–C, a Class of Diterpenoid Alkaloids with Unpredicted Conformations from <i>Croton cascarilloides</i>

Cascarinoids A–C (<b>1</b>–<b>3</b>), a new class of diterpenoid alkaloids with unpredicted conformations, were isolated and structurally characterized from <i>Croton cascarilloides</i>. It was demonstrated that the dispersion interaction might be one of the main contributors that stabilized the folded conformations for compounds <b>1</b>–<b>3</b>. Compounds <b>2</b> and <b>3</b> showed moderate immunosuppressive activity against T and/or B lymphocyte cells