The advancement of single-cell sequencing has progressed our ability to solve biological questions. Cell type annotation is of vital importance to this process, allowing for the analysis and interpretation of enormous single-cell datasets. At present, however, manual cell annotation which is the predominant approach remains limited by both speed and the requirement of expert knowledge.To address these challenges, we constructed a hierarchically organized marker map through manually curating over 26,000 cell marker entries from about 7000 publications. We then developed WISE, a weighted and integrated gene set enrichment method, to integrate the prevalence of canonical markers and ordered differentially expressed genes of specific cell types in the marker map. Benchmarking analysis suggested that our method outperformed state-of-the-art methods.By integrating the marker map and WISE, we developed a user-friendly and convenient web server, ACT ( http://xteam.xbio.top/ACT/ or http://biocc.hrbmu.edu.cn/ACT/ ), which only takes a simple list of upregulated genes as input and provides interactive hierarchy maps, together with well-designed charts and statistical information, to accelerate the assignment of cell identities and made the results comparable to expert manual annotation. Besides, a pan-tissue marker map was constructed to assist in cell assignments in less-studied tissues. Applying ACT to three case studies showed that all cell clusters were quickly and accurately annotated, and multi-level and more refined cell types were identified.We developed a knowledge-based resource and a corresponding method, together with an intuitive graphical web interface, for cell type annotation. We believe that ACT, emerging as a powerful tool for cell type annotation, would be widely used in single-cell research and considerably accelerate the process of cell type identification. 

Annotation of cell types (ACT): a convenient web server for cell type annotation

Decoding the basis of histological variation in human cancer.

Hydroperoxides (ROOH) are known as damaging agents capable of mediating mutation, while a role as signaling agents through oxidation of protein sulfhydryls that can alter cancer-related pathways has gained traction. Glutathione peroxidase 2 (GPX2) is an antioxidant enzyme that reduces ROOH at the expense of glutathione (GSH). GPX2 is noted for a tendency of large in-creases or decreases in expression levels during tumorigenesis that leads to investigators focusing on its role in cancer. However, GPX2 is only one component of multiple enzyme families that metabolize ROOH and GPX2 levels are often very low in the context of these other ROOH reducing activities. Colorectal cancer (CRC) was selected as a case study for examining GPX2 function as colorectal tissues and cancers are sites where GPX2 is highly expressed. A case can be made for a significant impact of changes in expression levels. There is also a link between GPX2 and NADPH oxidase 1 (NOX1) from earlier studies that is seldom addressed and is discussed, presenting data on a unique association in colon and CRC. Tumor-derived cell lines are quite commonly used for pre-clinical studies involving the role of GPX2 in CRC. Generally, selection for this type of work is limited to identifying cell lines based on high and low GPX2 expression with the standard re-search scheme of overexpression in low expressing lines and suppression in high expressing lines to identify impacted pathways. This overlooks CRC subtypes among cell lines involving a wide range of gene expression profiles and a variety of driver mutation differences along with a large difference in GPX2 expression levels. A trend for low and high GPX2 expressing cell lines to segregate into different CRC subclasses, indicated in this report, suggests that choices based solely on GPX2 levels may provide misleading and conflicting results by disregarding other properties of cell lines and failing to factor in differences in potential protein targets of ROOH. CRC and cell line classification schemes are presented here that were intended to assist workers in performing pre-clinical studies but are largely unnoted in work on GPX2 and CRC. Studies are often initiated on the premise that the transition from normal to CRC is associated with up-regulation of GPX2. This is probably correct. However, the source normal cells for CRC could be almost any colon cell type, some with very high GPX2 levels. These factors are addressed in this study. 

Evaluation of the Use of Cell Lines in Studies of Selenium-Dependent Glutathione Peroxidase 2 (GPX2) Involvement in Colorectal Cancer

Colorectal mucinous adenocarcinoma (MAC) is a particular pathological type that has yet to be thoroughly studied. This study aims to investigate the characteristics of colorectal MAC-related genes in colorectal cancer (CRC), explore the role of MAC-related genes in accurately classifying CRC, and further construct a prognostic signature.CRC samples were collected from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO). MAC-related differentially expressed genes (DEGs) were analyzed in TCGA samples. Based on colorectal MAC-related genes, TCGA CRC samples were molecularly typed by the non-negative matrix factorization (NMF). According to the molecular subtype characteristics, the RiskScore signature was constructed through univariate Cox, the least absolute shrinkage and selection operator (LASSO), and multivariate Cox regression analyses. Clinical significance in CRC of the RiskScore signature was analyzed. A nomogram was further built based on the RiskScore signature.From the colorectal MAC-related genes, three distinct molecular subtypes were identified. A RiskScore signature composed of six CRC subtype-related genes (CALB1, MMP1, HOXC6, ZIC2, SFTA2, and HYAL1) was constructed. Patients with high-RiskScores had the worse prognoses. RiskScores led to differences in gene mutation characteristics, antitumor drug sensitivity, and tumor microenvironment of CRC. A nomogram based on the signature was developed to predict the one-, three-, and five-year survival of CRC patients.MAC-related genes were able to classify CRC. A RiskScore signature based on the colorectal MAC-related molecular subtype was constructed, which had important clinical significance for guiding the accurate stratification of CRC patients. 

Identification and validation of a novel six-gene signature based on mucinous adenocarcinoma-related gene molecular typing in colorectal cancer

Colorectal cancer is a highly heterogeneous disease. Most colorectal cancers are classical adenocarcinoma, and mucinous adenocarcinoma is a unique histological subtype that is known to respond poorly to chemoradiotherapy. The difference in prognosis between mucinous adenocarcinoma and classical adenocarcinoma is controversial. Here, to gain insight into the differences between classical adenocarcinoma and mucinous adenocarcinoma, we analyse 7 surgical tumour samples from 4 classical adenocarcinoma and 3 mucinous adenocarcinoma patients by single-cell RNA sequencing. Our results indicate that mucinous adenocarcinoma cancer cells have goblet cell-like properties, and express high levels of goblet cell markers (REG4, SPINK4, FCGBP and MUC2) compared to classical adenocarcinoma cancer cells. TFF3 is essential for the transcriptional regulation of these molecules, and may cooperate with RPS4X to eventually lead to the mucinous adenocarcinoma mucus phenotype. The observed molecular characteristics may be critical in the specific biological behavior of mucinous adenocarcinoma. 

/pdf/single-cell-profiling-reveals-differences-between-human-u68on5wh.pdf

Single-cell profiling reveals differences between human classical adenocarcinoma and mucinous adenocarcinoma

BACKGROUND Surgery for thyroid carcinoma offers a good prognosis; however, cervical lymph node metastasis may occur in the early stage. An effective diagnostic method can accurately guide clinical surgical planning and the scope of lymph node dissection, ultimately improving patient prognosis. AIM To explore the diagnostic value of fine-needle aspiration of thyroglobulin (FNA-Tg) combined with ultrasound (US)-guided fine-needle aspiration cytology for cervical lymph node metastasis in thyroid carcinoma. METHODS We enrolled 209 pathologically confirmed thyroid carcinoma patients who visited our hospital between Jan 2017 and Dec 2020. Patients were tentatively diagnosed with cervical lymph node enlargement using preoperative US. They underwent US-guided fine-needle aspiration cytology and FNA-Tg. The value of single and combined application of the two methods for the diagnosis of cervical lymph node metastasis was calculated. The factors affecting FNA-Tg for diagnosis were analyzed using univariate and multivariate methods. RESULTS FNA-Tg values were significantly higher among patients with positive cervical lymph node metastasis. The sensitivity and specificity of US-guided fine-needle aspiration cytology, FNA-Tg, and US-guided fine-needle aspiration cytology + FNA-Tg were 85.48% and 90.59%, 83.06% and 87.06%, and 96.77% and 91.76%, respectively. The area under the receiver operating characteristic curve for US-guided fine-needle aspiration cytology, FNA-Tg, and the two combined, was 0.880, 0.851, and 0.943, respectively. A long diameter/short diameter ratio < 2, an insufficient number of acquired cells, a low serum thyroglobulin level, and an absence of typical metastatic US features increased the risk of cervical lymph node metastasis in thyroid carcinoma patients misdiagnosed using FNA-Tg. CONCLUSION The diagnostic value of FNA-Tg for detecting cervical lymph node metastasis is not high; however, combined with US-guided fine-needle aspiration cytology, it is significantly improved.

Value of thyroglobulin combined with ultrasound-guided fine-needle aspiration cytology for diagnosis of lymph node metastasis of thyroid carcinoma

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Single-cell profiling reveals differences between human classical adenocarcinoma and mucinous adenocarcinoma

Value of thyroglobulin combined with ultrasound-guided fine-needle aspiration cytology for diagnosis of lymph node metastasis of thyroid carcinoma