Single-Cell Profiling Reveals Heterogeneity of Primary and Lymph Node Metastatic Tumors and Immune Cell Populations and Discovers Important Prognostic Significance of CCDC43 in Oral Squamous Cell Carcinoma.

Although substantial progress has been made in biological research and clinical treatment in recent years, the clinical prognosis of oral squamous cell carcinoma (OSCC) is still not satisfactory. Tumor immune microenvironment (TIME) is a potential target, which plays an essential role in the response of anti-tumor immunity and immunotherapy. In this study, we used scRNA-seq data, revealing the heterogeneity of TIME between metastatic and primary site. We found that in the metastatic site, the content of cytotoxic T cells and classical activated macrophages (M1 macrophages) increases significantly, while alternately activated macrophages (M2 macrophages) and inflammatory cancer-associated fibroblasts (iCAFs) decrease, which may be due to the increased immunogenicity of OSCC cells in the metastatic site and the changes in some signal pathways. We also found that iCAFs may recruit alternately activated macrophages (M2 macrophages) by secreting CXCL12. Then, we described a regulatory network for communication between various TIME cells centered on OSCC cells, which can help to clarify the possible mechanism of lymph node metastasis in OSCC cells. By performing pseudotime trajectory analysis, we found that the expression CCDC43 is upregulated in more advanced OSCC cells and is an independent prognostic factor for poor living conditions. Other than this, the high expression of CCDC43 may impair the antitumor immunity of the human body and promote the metastasis of OSCC cells. Our research provides a profound insight into the immunological study of OSCC and an essential resource for future drug discovery.

Antitumor Effect of Saikosaponin A on Human Neuroblastoma Cells.

OBJECTIVE: Neuroblastoma (NB) is a highly metastatic tumor in children that develops in the sympathetic nervous system and has a low curative rate. Saikosaponin A (SSA), an active ingredient isolated from the root of Radix Bupleuri, is a natural compound with various pharmacological activities and shows good application prospects in antitumors. This study investigated the antihuman NB activity of SSA and underlying mechanisms associated with its actions. MATERIALS AND METHODS: The MTT method was used to detect the activity of SSA in inhibiting human NB cell SK-N-AS proliferation. Cell morphology was observed. The flow cytometry technology was used in analyzing the cell apoptosis rate. The Transwell assay evaluated cell migration and invasion following SSA treatment, apoptosis-related protein expression, and angiogenesis-related protein expression, and EMT-related proteins were detected by western blot analysis. RESULTS: SSA showed an inhibitory effect on SK-N-AS cells with the IC50 values of 14.14 µM at 24 h and 12.41 µM at 48 h. Results indicated that SSA has proapoptotic activity, and its proapoptotic activity is positively correlated with the Bax/Bcl-2/caspase-9/caspase-7/PARP pathway. Furthermore, SSA inhibited the invasion and migration of SK-N-AS cells via regulating the angiogenesis-related VEGFR2/Src/Akt pathway and the epithelial-mesenchymal transition- (EMT-) related protein expression. CONCLUSION: SSA exerts an antihuman NB effect and thus provides foundations for NB treatment.

Targeting tumor-associated macrophages in the tumor microenvironment.

Tumor-associated macrophages (TAMs) are the most abundant population type of tumor-infiltrating immune cells found in the tumor microenvironment (TME), and are evolutionarily associated with microvessel density in tumor tissues. TAMs can be broadly divided into M1-like and M2-like TAMs, which demonstrate antitumor and pro-tumor activity in the TME, respectively. Studies have indicated that: i) The predominate presence of M2-like TAMs in the TME can result in tumor immunosuppression and chemoresistance; ii) the ratio of M1-like to M2-like TAMs in the TME is positively correlated with better long-term prognosis of patients with cancer; iii) epigenetic silencing, preventing the secretion of M1-like TAM-associated molecules, is an important immune evasion mechanism during tumor progression; and iv) the transformation from M2-like to M1-like TAMs following exposure to specific conditions can result in tumor regression. The present study discusses the molecular events underlying the recruitment of macrophages and their polarization into M1-like or M2-like TAMs, and their differential roles in angiogenesis, angiostasis, invasion, metastasis and immune activity in the TME. This insight may inform the improved design of TAM-targeted cancer immunotherapy. Some of these therapeutic strategies show promising effects; however, challenges remain.

The cross-sectional study of hepatic lipase SNPs and plasma lipid levels.

By the combination of meta-analysis, the data of the 1,000 Genomes Project Phase 3, and the promoter sequence of hepatic lipase (LIPC), we performed the cross-sectional study to explore the associations of four variants (rs1077835; rs1077834; rs1800588 [C-514T], and rs2070895 [G-250A]) in LIPC promoter with plasma lipid levels. Our results indicate that the first and the next three of the four SNPs are, respectively, reported to be associated with the decreased and increased HDL-c level. Meta-analysis of 87 studies with 101,988 participants indicates that HDL-c level in rs1800588 (C-514T) (pooled mean difference = 0.03, 95%CI (0.03, 0.04), p < .001) and rs2070895 (G-250A) (pooled mean difference = 0.07, 95%CI (0.05, 0.09), p < .001) is higher in allele T or A carriers. Similarly, LDL-c, TC, TG, and BMI levels are generally increased in T or A alleles carriers. We failed to conduct the meta-analysis of rs1077835 and rs1077834 due to the limited previous reports. Data from the 1,000 Genomes indicate that the allele frequencies of the four SNPs in total or subpopulations are almost equal to each other. The paired value r 2 and D' of the four SNPs are larger than 0.8, which indicate the linkage disequilibrium of the four variants. The analysis of LIPC promoter indicate that C-514T and G-250A are, respectively, located in transcriptional factor binding sites of USF1and Pbx1b, which may partly explain the effect of the two SNPs on the decreased LIPC activity in the alleles carriers and the corresponding increased plasma lipids hydrolyzed by LIPC. These results may help us to better understand the different effects of the four SNPs on the plasma lipid levels among subpopulations and offer clues for future clinical treatment of dyslipidemia-related diseases.

A bibliometric analysis of long non-coding RNA and chemotherapeutic resistance research.

The global outputs of annual publication in long non-coding RNAs (lncRNAs) and chemotherapeutic resistance research exponentially increased from 2 in 2008 to 176 in 2017. Using Java application CiteSpace V and VOSviewer, this study assessed the publication model of lncRNAs and chemoresistance by bibliometric analysis. Totally, 2883 authors contributed 528 publications of lncRNAs and chemoresistance in 215 academic journals in the recent decade (2008-2018). Oncotarget in the 215 academic journals published the highest number of publications (60). China had the highest number of publication outputs (358). The leading institute was Nanjing Medical University. Wang Y was the most influential author (13 counts). Gupta RA had the most cited documents (87 counts). "Gene expression" and "poor prognosis" were identified as the hotspots. "Cancer stem cell", "HOTAIR" and "UCA1" were the frontiers of the fields in recent years. The increase of publications on lncRNAs and chemotherapeutic resistance will continue in the next years. HOTAIR and UCA1 with multiple roles in drug resistance may offer big opportunities for targeted chemoresistance in cancer therapy. These results may help us discover and explain the possible underlying laws of the subject.

Correction to: The imbalance in the complement system and its possible physiological mechanisms in patients with lung cancer.

Following publication of the original article [1], it was noticed that Fig. 3c was omitted from the final published article.

The imbalance in the complement system and its possible physiological mechanisms in patients with lung cancer.

BACKGROUND: The clinical and experimental evidences for complement-cancer relationships are solid, whereas an epidemiological study reporting the imbalance of complement system in patients is still lacking. METHODS: Using publicly available databases, we jointly compared the levels of complement components in plasma and lung cancer tissues. With iTRAQ proteomics, quantitative RT-PCR and western blotting, we analysed the differences in complement levels in lung cancer tissues and normal control tissues. Complement components are mainly synthesized by the liver and secreted into the blood. Using paired co-cultures of human normal QSG-7701 hepatocytes with lung cancer cells (A549, LTEP-α-2 or NCI-H1703) or human normal bronchial epithelial (HBE) cells, we examined the effects of lung cancer cells on complement synthesis and secretion in QSG-7701 hepatocytes. RESULTS: An integrated analysis of transcriptome and proteome datasets from 43 previous studies revealed lower mRNA and protein levels of 25 complement and complement-related components in lung cancer tissues than those in normal control tissues; conversely, higher levels of complement proteins were detected in plasma from patients than those in healthy subjects. Our iTRAQ proteome study identified decreased and increased levels of 31 and 2 complement and complement-related proteins, respectively, in lung cancer tissues, of which the reduced levels of 10 components were further confirmed using quantitative RT-PCR and western blotting. Paired co-cultures of QSG-7701 hepatocytes with A549, LTEP-α-2, NCI-H1703 or HBE cells indicated that lung cancer cells increased complement synthesis and secretion in QSG-7701 cells compared to HBE cells. CONCLUSIONS: The opposite associations between the levels of complement and complement-related components in lung cancer tissues and plasma from patients that have been repeatedly reported by independent publications may indicate the prevalence of an imbalance in the complement system of lung cancer patients. The possible mechanism of the imbalance may be associated not only with the decreased complement levels in lung cancer tissues but also the concurrent lung cancer tissue-induced increase in hepatocyte complement synthesis and plasma secretion in patients. And the imbalance should be accompanied by a suppression of complement-dependent immunity in lung cancer tissues coupled with a burden of complement immunity in the circulation of patients.

TdIF1: a putative oncogene in NSCLC tumor progression.

TdT-interacting factor 1 (TdIF1) is a ubiquitously expressed DNA- and protein-binding protein that directly binds to terminal deoxynucleotidyl transferase (TdT) polymerase. Little is known about the functional role of TdIF1 in cancer cellular signaling, nor has it previously been identified as aberrant in any type of cancer. We report here for the first time that TdIF1 is abundantly expressed in clinical lung cancer patients and that high expression of TdIF1 is associated with poor patient prognosis. We further established that TdIF1 is highly expressed in human non-small cell lung cancer (NSCLC) cell lines compared to a normal lung cell line. shRNA-mediated gene silencing of TdIF1 resulted in the suppression of proliferation and anchorage-independent colony formation of the A549 adenocarcinoma cell line. Moreover, when these TdIF1-silenced cells were used to establish a mouse xenograft model of human NSCLC, tumor size was greatly reduced. These data suggest that TdIF1 is a potent regulator of lung tumor development. Several cell cycle-related and tumor growth signaling pathways, including the p53 and HDAC1/2 pathways, were identified as participating in the TdIF1 signaling network by in silico analysis. Microarray, transcriptome and protein-level analyses validated p53 and HDAC1/2 modulation upon TdIF1 downregulation in an NSCLC cellular model. Moreover, several other cell cycle regulators were affected at the transcript level by TdIF1 silencing, including an increase in CDKN1A/p21 transcripts. Taken together, these results indicate that TdIF1 is a bona fide tumor-promoting factor in NSCLC and a potential target for therapy.

WeChat: An applicable and flexible social app software for mobile teaching.

The rapid development and popularization of smart phones and mobile internet has created a new social lifestyle, and correspondingly prompts the transformation of network teaching from desktop computer to mobile teaching. This article has compared the pros and cons of Tsinghua Education Online and WeChat official account (WOA) in fulfilling teaching functions. We also described the construction of WOA platform with the example of WOA-based teaching in biochemistry and molecular biology. The platform can establish nearly 75 menu catalogs and 2,250 items, which is capable for the publication of any types of teaching materials and information. The WOA teaching is well accepted and becomes popular in China due to the free, interactive, attractive, adaptable, portable, sustainable, and more participatory teaching styles. © 2018 International Union of Biochemistry and Molecular Biology, 46(5):555-560, 2018.

Synergetic and Antagonistic Molecular Effects Mediated by the Feedback Loop of p53 and JNK between Saikosaponin D and SP600125 on Lung Cancer A549 Cells.

We jointly analyzed the changes in cell cycle arrest and distribution, the accumulation of subphase cells, apoptosis, and proliferation in A549 cells treated with Saikosaponin D (Ssd) and JNK inhibitor SP600125 alone or in combination. Our results indicated that cell cycle arrest at G0/G1, S, and G2/M phases was coupled with the accumulation of subG1, subS, and subG2 cells, corresponding to early apoptosis, DNA endoreplication, and later inhibitory proliferation, respectively. Analyzing the expression of 18 cell cycle regulatory genes and JNK and phosphorylated JNK (pJNK) levels revealed an enhancement in these factors by Ssd. Additional SP600125 weakened or eliminated the Ssd-induced increase of these factors except that p53/p21 and Rassfia levels were further improved. Ingenuity Pathway Analysis (IPA) of the interactions of these factors revealed a negative synergistic effect on apoptosis while a positive synergistic effect on proliferative inhibition of the two drugs: (1) Ssd induced apoptosis via the activation of two axes, TGFα-JNK-p53 and TGFα-Rassfia-Mst1. By eliminating the Ssd-induced increase of JNK/pJNK, additional SP600125 weakened the Ssd-induced apoptotic axis of TGFα-JNK-p53 and simultaneously abolished Ssd-induced apoptosis; (2) Ssd inhibited proliferation by the activation of two axes, TGFß-p53/p21/p27/p15/p16 and TGFα-Rassfia-cyclin D1. By improving the Ssd-induced increase of p53/p21 and Rassfia, additional SP600125 enhanced the two axes of Ssd-induced inhibitory proliferation. Analyzing JNK/pJNK, p53, phospho-p53, and TNF-α levels revealed an opposite association of JNK/pJNK with p53 while consistent with phospho-p53 and TNF-α, which supported the proposals that JNK/pJNK negatively regulated p53 level, while it mediated p53 phosphorylation to transcriptionally activate TNF-α expression of apoptotic gene and trigger apoptosis. With the multiple roles, JNK/pJNK forms a synergetic and antagonistic feedback loop with phospho-p53/p53. Within the feedback loop, (1) Ssd-induced apoptosis depended on JNK/pJNK activities mediating phospho-p53 that activated TNF-α expression; (2) by weakening the negative regulation of JNK/pJNK in p53, SP600125 enhanced p53 level and the Ssd-induced inhibitory proliferation axes of TGFß-p53/p21/p27/p15/p16. The results indicated the central coordinating roles of the feedback loop in the synergistic and antagonistic effects of the two drugs in A549 cells and provided a rationale for the combination of Ssd with SP600125 in the treatment of lung cancer.

Evolutionary expansion of nematode-specific glycine-rich secreted peptides.

Genome-wide comparisons across 10 species from algae Guillardia theta to mammal human indicated that Caenorhabditis elegans and Caenorhabditis briggsae were highly enriched for glycine-rich secreted peptides (GRSPs) (110 GRSPs in C. elegans and 93 in C. briggsae) in this study. Chromosomal mapping showed that most GRSPs were clustered on the two nematode genomes [103 (93.64%) in C. elegans and 82 (88.17%) in C. briggsae], which could be divided into 18 cluster units in C. elegans and 13 in C. briggsae, respectively. Except for four C. elegans GRSPs clusters without matching clusters in C. briggsae, all other GRSPs clusters had paired synteny block between the two nematode genomes. Analyzing transcriptome datasets quantified by microarray indicated extensive genome-wide co-expression of GRSPs clusters after C. elegans infections. Highly homologous coding sequences and conserved exon-intron structures indicated that GRSPs tight clusters were likely derived from local DNA duplications. Phylogenetic conservation of synteny blocks between their genomes, co-expression of GRSPs clusters after C. elegans infections, and strong purifying selections of coding sequences may indicate evolutionary constraints acting on C. elegans to guarantee that C. elegans could mount rapid systematic responses to infections by co-expression, co-regulation, and co-functionality of GRSPs clusters.

Tudor domain-containing proteins of Drosophila melanogaster.

Tudor domain-containing proteins (Tudor proteins), which recognize and bind to methyl-arginine / lysine residues, play important roles in diverse epigenetics, gene expression and the regulation of various small RNAs. Using the complete set of 23 Tudor proteins from Drosophila, together with the available functional information, we propose a putative link for different types of Tudor domains (histone-binding, SMN and SND1) and the four functional groups of Tudor proteins (Group 1, binding the methyl-lysine / arginine of histone tails; Group 2, binding the methyl-RG / RA box of ligand; Group 3, binding the methyl-RG /RA box of microRNPs; and Group 4, binding the methyl-RG /RA box of PIWI proteins). Tudor domain types are distinguished by the nature of the sequence flanking the canonical Tudor domains. Sequence analysis indicates that Tudor domains experienced stepwise transit from one type to another during evolution. Tudor proteins of Group 4, collectively representing the great majority of Tudor proteins in Drosophila, are characterized by multiple Tudor domain repeats, which might be required for associating with several molecules of the same germ granule components. Tudor domain, a segment of approximately 60 amino acid residues, has been found in fungi, protozoa, unicellular eukaryota, plants and metazoa but not in the Guillardia theta nucleomorph. Similar frequencies of Tudor-containing genes (Tudor genes) among vertebrates and the frequent occurrence of orthologues among vertebrates, along with similar observations within arthropods suggest that Tudor genes are inherited largely vertically during evolution within different phylogenetic lineages.

Comprehensively surveying structure and function of RING domains from Drosophila melanogaster.

Using a complete set of RING domains from Drosophila melanogaster, all the solved RING domains and cocrystal structures of RING-containing ubiquitin-ligases (RING-E3) and ubiquitin-conjugating enzyme (E2) pairs, we analyzed RING domains structures from their primary to quarternary structures. The results showed that: i) putative orthologs of RING domains between Drosophila melanogaster and the human largely occur (118/139, 84.9%); ii) of the 118 orthologous pairs from Drosophila melanogaster and the human, 117 pairs (117/118, 99.2%) were found to retain entirely uniform domain architectures, only Iap2/Diap2 experienced evolutionary expansion of domain architecture; iii) 4 evolutionary structurally conserved regions (SCRs) are responsible for homologous folding of RING domains at the superfamily level; iv) besides the conserved Cys/His chelating zinc ions, 6 equivalent residues (4 hydrophobic and 2 polar residues) in the SCRs possess good-consensus and conservation- these 4 SCRs function in the structural positioning of 6 equivalent residues as determinants for RING-E3 catalysis; v) members of these RING proteins located nucleus, multiple subcellular compartments, membrane protein and mitochondrion are respectively 42 (42/139, 30.2%), 71 (71/139, 51.1%), 22 (22/139, 15.8%) and 4 (4/139, 2.9%); vi) CG15104 (Topors) and CG1134 (Mul1) in C3HC4, and CG3929 (Deltex) in C3H2C3 seem to display broader E2s binding profiles than other RING-E3s; vii) analyzing intermolecular interfaces of E2/RING-E3 complexes indicate that residues directly interacting with E2s are all from the SCRs in RING domains. Of the 6 residues, 2 hydrophobic ones contribute to constructing the conserved hydrophobic core, while the 2 hydrophobic and 2 polar residues directly participate in E2/RING-E3 interactions. Based on sequence and structural data, SCRs, conserved equivalent residues and features of intermolecular interfaces were extracted, highlighting the presence of a nucleus for RING domain fold and formation of catalytic core in which related residues and regions exhibit preferential evolutionary conservation.

Origin and evolution of ubiquitin-conjugating enzymes from Guillardia theta nucleomorph to hominoid.

The origin of eukaryotic ubiquitin-conjugating enzymes (E2s) can be traced back to the Guillardia theta nucleomorph about 2500 million years ago (Mya). E2s are largely vertically inherited over eukaryotic evolution [Lespinet, O., Wolf, Y.I., Koonin, E.V., Aravind, L., 2002. The role of lineage-specific gene family expansion in the evolution of eukaryotes. Genome Res. 1048-1059], while mammal E2s experienced evolution of multigene families by gene duplications which have been accompanied by the increase in the species complexity. Because of alternatively splicing, primate-specific expansions of E2s happened once again at a transcriptional level. Both of them resulted in increasing genomic complexity and diversity of primate E2 proteomic function. The evolutionary processes of human E2 gene structure during expansions were accompanied by exon duplication and exonization of intronic sequences. Exonizations of Transposable Elements (TEs) in UBE2D3, UBE2L3 and UBE2V1 genes from primates indicate that exaptation of TEs also plays important roles in the structural innovation of primate-specific E2s and may create alternative splicing isoforms at a transcriptional level. Estimates for the ratio of dN/dS suggest that a strong purifying selection had acted upon protein-coding sequences of their orthologous UBE2D2, UBE2A, UBE2N, UBE2I and Rbx1 genes from animals, plants and fungi. The similar rates of synonymous substitutions are in accordance with the neutral mutation-random drift hypothesis of molecular evolution. Systematic detection of the origin and evolution of E2s, analyzing the evolution of E2 multigene families by gene duplications and the evolutionary processes of E2s during expansions, and testing its evolutionary force using E2s from distant phylogenetic lineages may advance our distinguishing of ancestral E2s from created E2s, and reveal previously unknown relationships between E2s and metazoan complexity. Analysis of these conserved proteins provides strong support for a close relationship between social amoeba and eukaryote, choanoflagellate and metazoan, and for the central roles of social amoeba and choanoflagellate in the origin and evolution of eukaryote and metazoan. Retracing the different stages of primate E2 exonization by monitoring genomic events over 63 Myr of primate evolution will advance our understanding of how TEs dynamically modified primate transcriptome and proteome in the past, and continue to do so.