Specific RNA m6A modification sites in bone marrow mesenchymal stem cells from the jawbone marrow of type 2 diabetes patients with dental implant failure / 国际口腔科学杂志·英文版

The failure rate of dental implantation in patients with well-controlled type 2 diabetes mellitus (T2DM) is higher than that in non-diabetic patients. This due, in part, to the impaired function of bone marrow mesenchymal stem cells (BMSCs) from the jawbone marrow of T2DM patients (DM-BMSCs), limiting implant osseointegration. RNA N6-methyladenine (m6A) is important for BMSC function and diabetes regulation. However, it remains unclear how to best regulate m6A modifications in DM-BMSCs to enhance function. Based on the "m6A site methylation stoichiometry" of m6A single nucleotide arrays, we identified 834 differential m6A-methylated genes in DM-BMSCs compared with normal-BMSCs (N-BMSCs), including 43 and 790 m6A hypermethylated and hypomethylated genes, respectively, and 1 gene containing hyper- and hypomethylated m6A sites. Differential m6A hypermethylated sites were primarily distributed in the coding sequence, while hypomethylated sites were mainly in the 3'-untranslated region. The largest and smallest proportions of m6A-methylated genes were on chromosome 1 and 21, respectively. MazF-PCR and real-time RT-PCR results for the validation of erythrocyte membrane protein band 4.1 like 3, activity-dependent neuroprotector homeobox (ADNP), growth differentiation factor 11 (GDF11), and regulator of G protein signalling 2 agree with m6A single nucleotide array results; ADNP and GDF11 mRNA expression decreased in DM-BMSCs. Furthermore, gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses suggested that most of these genes were enriched in metabolic processes. This study reveals the differential m6A sites of DM-BMSCs compared with N-BMSCs and identifies candidate target genes to enhance BMSC function and improve implantation success in T2DM patients.

Promising applications of human-derived saliva biomarker testing in clinical diagnostics / 国际口腔科学杂志·英文版

Saliva testing is a vital method for clinical applications, for its noninvasive features, richness in substances, and the huge amount. Due to its direct anatomical connection with oral, digestive, and endocrine systems, clinical usage of saliva testing for these diseases is promising. Furthermore, for other diseases that seeming to have no correlations with saliva, such as neurodegenerative diseases and psychological diseases, researchers also reckon saliva informative. Tremendous papers are being produced in this field. Updated summaries of recent literature give newcomers a shortcut to have a grasp of this topic. Here, we focused on recent research about saliva biomarkers that are derived from humans, not from other organisms. The review mostly addresses the proceedings from 2016 to 2022, to shed light on the promising usage of saliva testing in clinical diagnostics. We recap the recent advances following the category of different types of biomarkers, such as intracellular DNA, RNA, proteins and intercellular exosomes, cell-free DNA, to give a comprehensive impression of saliva biomarker testing.

Circular RNA circRSF1 binds to HuR to promote radiation-induced inflammatory phenotype in hepatic stellate cells / 南方医科大学学报

OBJECTIVE@#To investigate whether circular RNA circRSF1 regulates radiation-induced inflammatory phenotype of hepatic stellate cells (HSCs) by binding to HuR protein and repressing its function.@*METHODS@#Human HSC cell line LX2 with HuR overexpression or knockdown was exposed to 8 Gy X-ray irradiation, and the changes in the expression of inflammatory factors (IL-1β, IL-6 and TNF-α) were detected by qRT-PCR. The expressions of IκBα and phosphorylation of NF-κB were detected with Western blotting. The binding of circRSF1 to HuR was verified by RNA pull-down assay and RNA-binding protein immunoprecipitation (RIP). The expressions of inflammatory factors, IκBα and the phosphorylation of NF-κB were detected after modifying the interaction between circRSF1 and HuR.@*RESULTS@#Knockdown of HuR significantly up- regulated the expressions of IL-1β, IL-6 and TNF-α, decreased IκBα expression and promoted NF-κB phosphorylation in irradiated LX2 cells, whereas overexpression of HuR produced the opposite changes (P < 0.05). Overexpression or knockdown of circRSF1 did not significantly affect the expression of HuR. RNA pull-down and RIP experiments confirmed the binding between circRSF1 and HuR. Overexpression of circRSF1 significantly reduced the binding of HuR to IκBα and down-regulated the expression of IκBα (P < 0.05). Overexpression of circRSF1 combined with HuR overexpression partially reversed the up-regulation of the inflammatory factors, down-regulated IκBα expression and increased phosphorylation of NFκB in LX2 cells, while the opposite effects were observed in cells with knockdown of both circRSF1 and HuR (P < 0.05).@*CONCLUSION@#circRSF1 reduces IκBα expression by binding to HuR to promote the activation of NF-κB pathway, thereby enhancing radiation- induced inflammatory phenotype of HSCs.

Advances in the RNA-targeting CRISPR-Cas systems / 生物工程学报

The CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR associated proteins) system is an adaptive immune system of bacteria and archaea against phages, plasmids and other exogenous genetic materials. The system uses a special RNA (CRISPR RNA, crRNA) guided endonuclease to cut the exogenous genetic materials complementary to crRNA, thus blocking the infection of exogenous nucleic acid. According to the composition of the effector complex, CRISPR-Cas system can be divided into two categories: class 1 (including type Ⅰ, Ⅳ, and Ⅲ) and class 2 (including type Ⅱ, Ⅴ, and Ⅵ). Several CRISPR-Cas systems have been found to have very strong ability to specifically target RNA editing, such as type Ⅵ CRISPR-Cas13 system and type Ⅲ CRISPR-Cas7-11 system. Recently, several systems have been widely used in the field of RNA editing, making them a powerful tool for gene editing. Understanding the composition, structure, molecular mechanism and potential application of RNA-targeting CRISPR-Cas systems will facilitate the mechanistic research of this system and provide new ideas for developing gene editing tools.

Research progress on regulation of N6-adenylate methylation modification in lipid metabolism disorders / 生理学报

Lipid metabolism is a complex physiological process, which is closely related to nutrient regulation, hormone balance and endocrine function. It involves the interactions of multiple factors and signal transduction pathways. Lipid metabolism disorder is one of the main mechanisms to induce a variety of diseases, such as obesity, diabetes, non-alcoholic fatty liver disease, hepatitis, hepatocellular carcinoma and their complications. At present, more and more studies have found that the "dynamic modification" of N6-adenylate methylation (m6A) on RNA represents a new "post-transcriptional" regulation mode. m6A methylation modification can occur in mRNA, tRNA, ncRNA, etc. Its abnormal modification can regulate gene expression changes and alternative splicing events. Many latest references have reported that m6A RNA modification is involved in the epigenetic regulation of lipid metabolism disorder. Based on the major diseases induced by lipid metabolism disorders, we reviewed the regulatory roles of m6A modification in the occurrence and development of those diseases. These overall findings inform further in-depth investigations of the underlying molecular mechanisms regarding the pathogenesis of lipid metabolism disorders from the perspective of epigenetics, and provide reference for health prevention, molecular diagnosis and treatment of related diseases.

H19 recruited N 6 -methyladenosine (m 6 A) reader YTHDF1 to promote SCARB1 translation and facilitate angiogenesis in gastric cancer / 中华医学杂志(英文版)

BACKGROUND@#Angiogenesis is described as a complex process in which new microvessels sprout from endothelial cells of existing vasculature. This study aimed to determine whether long non-coding RNA (lncRNA) H19 induced the angiogenesis of gastric cancer (GC) and its possible mechanism.@*METHODS@#Gene expression level was determined by quantitative real-time polymerase chain reaction and western blotting. Cell counting kit-8, transwell, 5-Ethynyl-2'-deoxyuridine (EdU), colony formation assay, and human umbilical vein endothelial cells (HUVECs) angiogenesis assay as well as Matrigel plug assay were conducted to study the proliferation, migration, and angiogenesis of GC in vitro and in vivo . The binding protein of H19 was found by RNA pull-down and RNA Immunoprecipitation (RIP). High-throughput sequencing was performed and next Gene Ontology (GO) as well as Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis was conducted to analyze the genes that are under H19 regulation. Methylated RIP (me-RIP) assay was used to investigate the sites and abundance among target mRNA. The transcription factor acted as upstream of H19 was determined through chromatin immunoprecipitation (ChIP) and luciferase assay.@*RESULTS@#In this study, we found that hypoxia-induced factor (HIF)-1α could bind to the promoter region of H19, leading to H19 overexpression. High expression of H19 was correlated with angiogenesis in GC, and H19 knocking down could inhibit cell proliferation, migration and angiogenesis. Mechanistically, the oncogenic role of H19 was achieved by binding with the N 6 -methyladenosine (m 6 A) reader YTH domain-containing family protein 1 (YTHDF1), which could recognize the m 6 A site on the 3'-untransated regions (3'-UTR) of scavenger receptor class B member 1 (SCARB1) mRNA, resulting in over-translation of SCARB1 and thus promoting the proliferation, migration, and angiogenesis of GC cells.@*CONCLUSION@#HIF-1α induced overexpression of H19 via binding with the promoter of H19, and H19 promoted GC cells proliferation, migration and angiogenesis through YTHDF1/SCARB1, which might be a beneficial target for antiangiogenic therapy for GC.

TRIM25 inhibits HBV replication by promoting HBx degradation and the RIG-I-mediated pgRNA recognition / 中华医学杂志(英文版)

BACKGROUND@#The hepatitis B virus (HBV) vaccine has been efficiently used for decades. However, hepatocellular carcinoma caused by HBV is still prevalent globally. We previously reported that interferon (IFN)-induced tripartite motif-containing 25 (TRIM25) inhibited HBV replication by increasing the IFN expression, and this study aimed to further clarify the anti-HBV mechanism of TRIM25.@*METHODS@#The TRIM25-mediated degradation of hepatitis B virus X (HBx) protein was determined by detecting the expression of HBx in TRIM25-overexpressed or knocked-out HepG2 or HepG2-NTCP cells via Western blotting. Co-immunoprecipitation was performed to confirm the interaction between TRIM25 and HBx, and colocalization of TRIM25 and HBx was identified via immunofluorescence; HBV e-antigen and HBV surface antigen were qualified by using an enzyme-linked immunosorbent assay (ELISA) kit from Kehua Biotech. TRIM25 mRNA, pregenomic RNA (pgRNA), and HBV DNA were detected by quantitative real-time polymerase chain reaction. The retinoic acid-inducible gene I (RIG-I) and pgRNA interaction was verified by RNA-binding protein immunoprecipitation assay.@*RESULTS@#We found that TRIM25 promoted HBx degradation, and confirmed that TRIM25 could enhance the K90-site ubiquitination of HBx as well as promote HBx degradation by the proteasome pathway. Interestingly, apart from the Really Interesting New Gene (RING) domain, the SPRY domain of TRIM25 was also indispensable for HBx degradation. In addition, we found that the expression of TRIM25 increased the recognition of HBV pgRNA by interacting with RIG-I, which further increased the IFN production, and SPRY, but not the RING domain is critical in this process.@*CONCLUSIONS@#The study found that TRIM25 interacted with HBx and promoted HBx-K90-site ubiquitination, which led to HBx degradation. On the other hand, TRIM25 may function as an adaptor, which enhanced the recognition of pgRNA by RIG-I, thereby further promoting IFN production. Our study can contribute to a better understanding of host-virus interaction.

Rapid detection and genotyping of SARS-CoV-2 Omicron BA.4/5 variants using a RT-PCR and CRISPR-Cas12a-based assay / 南方医科大学学报

OBJECTIVE@#To establish a rapid detection and genotyping method for SARS-CoV-2 Omicron BA.4/5 variants using CRISPPR-Cas12a gene editing technology.@*METHODS@#We combined reverse transcription-polymerase chain reaction (RT-PCR) and CRISPR gene editing technology and designed a specific CRISPPR RNA (crRNA) with suboptimal protospacer adjacent motifs (PAM) for rapid detection and genotyping of SARS- CoV-2 Omicron BA.4/5 variants. The performance of this RT- PCR/ CRISPPR-Cas12a assay was evaluated using 43 clinical samples of patients infected by wild-type SARS-CoV-2 and the Alpha, Beta, Delta, Omicron BA. 1 and BA. 4/5 variants and 20 SARS- CoV- 2-negative clinical samples infected with 11 respiratory pathogens. With Sanger sequencing method as the gold standard, the specificity, sensitivity, concordance (Kappa) and area under the ROC curve (AUC) of RT-PCR/CRISPPR-Cas12a assay were calculated.@*RESULTS@#This assay was capable of rapid and specific detection of SARS- CoV-2 Omicron BA.4/5 variant within 30 min with the lowest detection limit of 10 copies/μL, and no cross-reaction was observed in SARS-CoV-2-negative clinical samples infected with 11 common respiratory pathogens. The two Omicron BA.4/5 specific crRNAs (crRNA-1 and crRNA-2) allowed the assay to accurately distinguish Omicron BA.4/5 from BA.1 sublineage and other major SARS-CoV-2 variants of concern. For detection of SARS-CoV-2 Omicron BA.4/5 variants, the sensitivity of the established assay using crRNA-1 and crRNA-2 was 97.83% and 100% with specificity of 100% and AUC of 0.998 and 1.000, respectively, and their concordance rate with Sanger sequencing method was 92.83% and 96.41%, respectively.@*CONCLUSION@#By combining RT-PCR and CRISPPR-Cas12a gene editing technology, we successfully developed a new method for rapid detection and identification of SARS-CoV-2 Omicron BA.4/5 variants with a high sensitivity, specificity and reproducibility, which allows rapid detection and genotyping of SARS- CoV-2 variants and monitoring of the emerging variants and their dissemination.

Effects of titanium dioxide nanoparticles on circRNA expression profiles in human hepatocellular carcinoma cells HepG2 / 北京大学学报(医学版)

OBJECTIVE@#To investigate the effect of titanium dioxide nanoparticles (TiO2 NPs) on the expression profile of circular ribonucleic acid (circRNA) in human hepatocytes through in vitro cell experiments, and to attempt to understand the potential mechanism of hepatotoxicity through bioinformatics analysis.@*METHODS@#TiO2 NPs were characterized from the aspects of particle size, shape and agglomeration state. The cell counting kit-8 (CCK8) was used to detect the cytotoxicity of TiO2 NPs against human hepatocellular carcinoma cells (HepG2) after exposure to 0, 1.56, 3.13, 6.25, 12.5, 25, 50, 100, and 200 mg/L TiO2 NPs for 24 h or 48 h. The cells were treated at doses of 0 mg/L TiO2 NPs (control group) and 100 mg/L TiO2 NPs (treatment group), and collected after exposure for 48 h, and then RNA from the extracted cell samples was collected and sequenced. The differential circRNAs between the control and the TiO2 NPs treatment groups were screened, and then the enrichment pathway of the differential circRNA target gene was analyzed by multivariate statistics. According to the sequencing results, significantly altered genes and important genes in the significant enrichment pathways were screened, and real-time reverse transcription-polymerase chain reaction (real-time RT-PCR) was performed to verify the results.@*RESULTS@#TiO2 NPs were spherical anatase with a hydrated particle size of (323.50±85.44) nm and a Zeta potential of (-21.00±0.72) mV in a serum-free medium. The results of the CCK8 cytotoxicity assay showed that with the increase of TiO2 NPs concentration, cell viability gradually decreased. A total of 11 478 circRNAs were found by RNA sequencing. Compared with the control groups, TiO2 NPs treatment groups (100 mg/L) had a total of 89 differential circRNAs, of which 59 were up-regulated and 30 were down-regulated. Analysis of the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway showed that the targeted genes of differential circRNAs were mainly enriched in fatty acid degradation, Fanconi anemia pathway, and fatty acid metabolism. The expression levels of circRNA.6730, circRNA.3650 and circRNA.4321 were significantly different between the TiO2 NPs treatment group and the control group, which were consistent with the sequencing results.@*CONCLUSION@#TiO2 NPs can induce changes in circRNA expression profile, and epigenetics may play an important role in the mechanism of hepatotoxicity.

Effects of RNA M6A demethylase ALKBH5 gene deficiency on morphology and function of cerebellum in aged mice / 中华病理学杂志

Objective: To investigate the effects of RNA m6A demethylase ALKBH5 gene deficiency on cerebellar morphology and function in the aged mice, and to explore the role of ALKBH5 in cerebellar degeneration. Methods: Western blot was performed to detect the protein level of ALKBH5 in the cerebellum of wild-type mice of various ages. The expression of NeuN, Calbindin-D28K, MAP2, GFAP and other proteins in the cerebella of middle-aged (12-month-old) and aged (18-month-old) wild-type mice and ALKBH5-/- mice was examined using immunohistochemistry. The balance beam test and gait analysis were performed to test the balance ability and motor coordination of the mice. Results: With aging of the mice, the expression of ALKBH5 in the cerebellum increased gradually in an age-dependent manner. In the aged mice, but not middle-aged mice, the body weight, whole brain weight and cerebellum weight of ALKBH5-/- mice decreased by 15%, 10% and 21%, respectively (P<0.05). The expression of ALKBH5 in the Purkinje cells was much higher than that in other types of neural cells. Correspondingly, ALKBH5-deficiency caused 40% reduction in the number of Purkinje cells, as well as the length and density of neuronal dendrites in the aged mice (P<0.01). In addition, the time for the aged ALKBH5-/- mice to pass the balance beam was 70% longer than that of the wild type mice of the same age, with unstable gaits (P<0.01). Conclusions: Gene deficiency of RNA m6A demethylase ALKBH5 causes cerebellar atrophy, Purkinje neuron loss and damage in the aged mice. These changes eventually affect mice's motor coordination and balance ability. These results suggest that imbalanced RNA m6A methylation may lead to neurodegenerative lesions in the cerebellum of mice.

Circular RNA expression profiles and circRNA-miRNA-mRNA crosstalk in pre-eclamptic placenta / 中华妇产科杂志

Objective: To identify the expression profile of circular RNA (circRNA) in placenta of pre-eclampsia (PE) pregnant women by high-throughput sequencing, and to construct the circRNA-microRNA (miRNA)-messenger RNA (mRNA) interaction network, so as to reveal the related pathways and regulatory mechanisms of PE. Methods: The clinical data and placentas of 42 women with PE (PE group) and 30 normal pregnant women (control group) who delivered in West China Second University Hospital from November 2019 to June 2021 were collected. (1) High-throughput sequencing was used to establish the differentially expressed circRNA profiles in placental tissues of 5 pairs of PE group and the control group. (2) Real-time quantitative PCR (qRT-PCR) was used to verify the expression levels of 6 differentially expressed circRNAs in placental tissues of PE group and control group. (3) Bioinformatics analysis was used to predict the target miRNA and analyze the co-expressed mRNA to construct a competitive endogenous RNA (ceRNA) network. The differentially expressed circRNAs were analyzed by Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathways. (4) Logistic regression analysis, Pearson correlation and Kendall's tau-b correlation analysis were used to test the correlation between the three differentially expressed circRNAs and the risk of PE and clinical characteristics. (5) circRNA_05393 was selected for subsequent functional study. Small interfering RNA (siRNA) and overexpression plasmid were used to knock down or increase the expression level of circRNA_05393 in trophoblast cell line HTR-8/SVneo cells, respectively. Transwell assay was used to detect the migration and invasion ability of the trophoblasts in vitro. Cell counting kit-8 assay was used to detect the proliferation ability of the trophoblasts. Results: (1) Seventy-two differentially expressed circRNAs were identified by high-throughput sequencing, of which 35 were up-regulated and 37 were down-regulated. (2) qRT-PCR showed that compared with the control group, circRNA_00673 (1.306±0.168 vs 2.059±0.242; t=2.356, P=0.021) and circRNA_07796 (1.275±0.232 vs 1.954±0.230; t=2.018, P=0.047) were significantly increased, while circRNA_05393 (1.846±0.377 vs 0.790±0.094; t=3.138, P=0.002) was significantly decreased. (3) The circRNA-miRNA-mRNA interaction network contained 3 circRNAs, 8 miRNAs and 53 mRNAs. GO functional annotation analysis showed that the biological process was mainly enriched in iron ion homeostasis, membrane depolarization during action potential and neuronal action potential. In terms of cellular components, they were mainly enriched in cytoskeleton and membrane components. In terms of molecular function, they were mainly enriched in the activity of voltage-gated sodium channel and basic amino acid transmembrane transporter. KEGG pathway enrichment analysis showed that mRNAs in the interaction network were mainly enriched in complement and coagulation cascade, glycine, serine and threonine metabolism, p53 signaling pathway and peroxisome proliferators-activated receptors (PPAR) signaling pathway. (4) Logistic regression analysis showed that down-regulation of circRNA_05393 expression was a risk factor for PE (OR=0.044, 95%CI: 0.003-0.596; P=0.019). Correlation analysis showed that circRNA_05393 was significantly correlated with systolic blood pressure and diastolic blood pressure in PE pregnant women (both P<0.05). (5) Knock down or overexpression of circRNA_05393 significantly reduced or increased the migration and invasion abilities of HTR-8/SVneo cells (all P<0.05), but had no significant effect on the ability of tube formation and proliferation (all P>0.05). Conclusions: The construction of circRNA expression profile in placenta and the exploration of circRNA-miRNA-mRNA interaction network provide the possibility to reveal the regulatory mechanism of specific circRNA involved in PE. Inhibition of circRNA_05393 may induce the progression of PE by reducing the migration and invasion of trophoblasts.

Research progress of m6A methylation in prostate cancer / 亚洲男科学杂志(英文版)

N6-methyladenosine (m6A) is a ubiquitous RNA modification in mammals. This modification is "written" by methyltransferases and then "read" by m6A-binding proteins, followed by a series of regulation, such as alternative splicing, translation, RNA stability, and RNA translocation. At last, the modification is "erased" by demethylases. m6A modification is essential for normal physiological processes in mammals and is also a very important epigenetic modification in the development of cancer. In recent years, cancer-related m6A regulation has been widely studied, and various mechanisms of m6A regulation in cancer have also been recognized. In this review, we summarize the changes of m6A modification in prostate cancer and discuss the effect of m6A regulation on prostate cancer progression, aiming to profile the potential relevance between m6A regulation and prostate cancer development. Intensive studies on m6A regulation in prostate cancer may uncover the potential role of m6A methylation in the cancer diagnosis and cancer therapy.

Role of N6-methyladenosine RNA methylation in central nervous system: a review / 生物工程学报

There are a variety of post-transcriptional modifications in mRNA, which regulate the stability, splicing, translation, transport and other processes of mRNA, followed by affecting cell development, body immunity, learning and cognition and other important physiological functions. m6A modification is one of the most abundant post-transcriptional modifications widely existing in mRNA, regulating the metabolic activities of RNA and affecting gene expression. m6A modified homeostasis is critical for the development and maintenance of the nervous system. In recent years, m6A modification has been found in neurodegenerative diseases, mental diseases and brain tumors. This review summarizes the role of m6A methylation modification in the development, function and related diseases of the central nervous system in recent years, providing potential clinical therapeutic targets for neurological diseases.

5'-tiRNA-Gln inhibits hepatocellular carcinoma progression by repressing translation through the interaction with eukaryotic initiation factor 4A-I / 医学前沿

tRNA-derived small RNAs (tsRNAs) are novel non-coding RNAs that are involved in the occurrence and progression of diverse diseases. However, their exact presence and function in hepatocellular carcinoma (HCC) remain unclear. Here, differentially expressed tsRNAs in HCC were profiled. A novel tsRNA, tRNAGln-TTG derived 5'-tiRNA-Gln, is significantly downregulated, and its expression level is correlated with progression in patients. In HCC cells, 5'-tiRNA-Gln overexpression impaired the proliferation, migration, and invasion in vitro and in vivo, while 5'-tiRNA-Gln knockdown yielded opposite results. 5'-tiRNA-Gln exerted its function by binding eukaryotic initiation factor 4A-I (EIF4A1), which unwinds complex RNA secondary structures during translation initiation, causing the partial inhibition of translation. The suppressed downregulated proteins include ARAF, MEK1/2 and STAT3, causing the impaired signaling pathway related to HCC progression. Furthermore, based on the construction of a mutant 5'-tiRNA-Gln, the sequence of forming intramolecular G-quadruplex structure is crucial for 5'-tiRNA-Gln to strongly bind EIF4A1 and repress translation. Clinically, 5'-tiRNA-Gln expression level is negatively correlated with ARAF, MEK1/2, and STAT3 in HCC tissues. Collectively, these findings reveal that 5'-tiRJNA-Gln interacts with EIF4A1 to reduce related mRNA binding through the intramolecular G-quadruplex structure, and this process partially inhibits translation and HCC progression.

Promising roles of non-exosomal and exosomal non-coding RNAs in the regulatory mechanism and as diagnostic biomarkers in myocardial infarction / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Non-exosomal non-coding RNAs (non-exo-ncRNAs) and exosomal ncRNAs (exo-ncRNAs) have been associated with the pathological development of myocardial infarction (MI). Accordingly, this analytical review provides an overview of current MI studies on the role of plasma non-exo/exo-ncRNAs. We summarize the features and crucial roles of ncRNAs and reveal their novel biological correlations via bioinformatics analysis. The following contributions are made: (1) we comprehensively describe the expression profile, competing endogenous RNA (ceRNA) network, and "pre-necrotic" biomarkers of non-exo/exo-ncRNAs for MI; (2) functional enrichment analysis indicates that the target genes of ncRNAs are enriched in the regulation of apoptotic signaling pathway and cellular response to chemical stress, etc.; (3) we propose an updated and comprehensive view on the mechanisms, pathophysiology, and biomarker roles of non-exo/exo-ncRNAs in MI, thereby providing a theoretical basis for the clinical management of MI.

Biomarcadores convencionales y emergentes en hepatitis B / Conventional and emerging biomarkers in hepatitis B

A nivel mundial, 300 millones de personas están infectadas por el virus de la hepatitis B (VHB). A pesar de que existe una vacuna que previene la infección y se dispone de tratamiento antiviral que suprime la replicación del virus, no hay cura aún. El principal problema que evita la recuperación total del paciente, incluso para aquel que recibe tratamiento, es la persistencia de dos formas del genoma viral en los hepatocitos: el ADN circular covalentemente cerrado (ADNccc), el cual se encuentra en forma de episoma y tiene la capacidad de replicarse, y las secuencias lineales subge-nómicas que se integran en el genoma humano, con potencial oncogénico. Hasta el momento se dispone de unos pocos biomarcadores para monitorear o predecir la progresión de la enfermedad y la respuesta al tratamiento. Estos biomarcadores se detectan durante la infección, y son la base para la monitorización de la enfermedad y hacer un diagnóstico de la fase clínica de la infección. Recientemente han surgido nuevos biomarcadores como el antígeno relacionado con el core del virus de la hepatitis B (HBcrAg) y la detección del ARN del VHB, que parecen correlacionarse con los niveles transcripcionales del ADNccc, además, durante el tratamiento parecen ayudar a predecir la respuesta y podrían identificar aquellos a quienes se les puede suspender la terapia sin riesgo de recaída. En esta revisión, se describe la utilidad de los principales biomarcadores convencionales en hepatitis B, y se abordan los dos biomarcadores emergentes más estudiados que prometen evaluar el curso de la infección, al igual que determinar la progresión de la enfermedad y la respuesta al tratamiento.

Globally, 300 million people are infected with hepatitis B virus (HBV). Although there is a vaccine that prevents infection and antiviral treatment that suppresses the replication of the virus, there is still no cure. The main problem that prevents the total recovery of the patient, even for those who recei-ve treatment, is the persistence of two forms of the viral genome in hepatocytes: covalently close circular DNA (cccDNA), which is in the form of an episome that has the ability to replicate, and linear subgenomic sequences that are integrated into the human genome, with oncogenic potential. Few biomarkers are currently available to monitor or predict disease progression and response to treatment. These biomarkers are detected during infection and are the basis for monitoring the di-sease and making a diagnosis of the clinical phase of the infection. New biomarkers have recently emerged, such as hepatitis B core-related antigen (HBcrAg) and HBV RNA detection, which seem to correlate with cccDNA transcriptional levels while during treatment seem to help predict response, and could identify those for whom therapy can be discontinued without risk of relapse. In this review, the usefulness of the main conventional biomarkers in hepatitis B is described, and the two most studied emerging biomarkers are mentioned, which promise to evaluate the course of the infection, as well as to determine disease progression and treatment response.

Extracellular signals regulate the biogenesis of extracellular vesicles

Extracellular vesicles (EVs) are naturally released membrane vesicles that act as carriers of proteins and RNAs for intercellular communication. With various biomolecules and specific ligands, EV has represented a novel form of information transfer, which possesses extremely outstanding efficiency and specificity compared to the classical signal transduction. In addition, EV has extended the concept of signal transduction to intercellular aspect by working as the collection of extracellular information. Therefore, the functions of EVs have been extensively characterized and EVs exhibit an exciting prospect for clinical applications. However, the biogenesis of EVs and, in particular, the regulation of this process by extracellular signals, which are essential to conduct further studies and support optimal utility, remain unclear. Here, we review the current understanding of the biogenesis of EVs, focus on the regulation of this process by extracellular signals and discuss their therapeutic value.

PathogenTrack and Yeskit: tools for identifying intracellular pathogens from single-cell RNA-sequencing datasets as illustrated by application to COVID-19 / 医学前沿

Pathogenic microbes can induce cellular dysfunction, immune response, and cause infectious disease and other diseases including cancers. However, the cellular distributions of pathogens and their impact on host cells remain rarely explored due to the limited methods. Taking advantage of single-cell RNA-sequencing (scRNA-seq) analysis, we can assess the transcriptomic features at the single-cell level. Still, the tools used to interpret pathogens (such as viruses, bacteria, and fungi) at the single-cell level remain to be explored. Here, we introduced PathogenTrack, a python-based computational pipeline that uses unmapped scRNA-seq data to identify intracellular pathogens at the single-cell level. In addition, we established an R package named Yeskit to import, integrate, analyze, and interpret pathogen abundance and transcriptomic features in host cells. Robustness of these tools has been tested on various real and simulated scRNA-seq datasets. PathogenTrack is competitive to the state-of-the-art tools such as Viral-Track, and the first tools for identifying bacteria at the single-cell level. Using the raw data of bronchoalveolar lavage fluid samples (BALF) from COVID-19 patients in the SRA database, we found the SARS-CoV-2 virus exists in multiple cell types including epithelial cells and macrophages. SARS-CoV-2-positive neutrophils showed increased expression of genes related to type I interferon pathway and antigen presenting module. Additionally, we observed the Haemophilus parahaemolyticus in some macrophage and epithelial cells, indicating a co-infection of the bacterium in some severe cases of COVID-19. The PathogenTrack pipeline and the Yeskit package are publicly available at GitHub.

METTL3-mediated m6A RNA methylation regulates dorsal lingual epithelium homeostasis / 国际口腔科学杂志·英文版

The dorsal lingual epithelium, which is composed of taste buds and keratinocytes differentiated from K14+ basal cells, discriminates taste compounds and maintains the epithelial barrier. N6-methyladenosine (m6A) is the most abundant mRNA modification in eukaryotic cells. How METTL3-mediated m6A modification regulates K14+ basal cell fate during dorsal lingual epithelium formation and regeneration remains unclear. Here we show knockout of Mettl3 in K14+ cells reduced the taste buds and enhanced keratinocytes. Deletion of Mettl3 led to increased basal cell proliferation and decreased cell division in taste buds. Conditional Mettl3 knock-in mice showed little impact on taste buds or keratinization, but displayed increased proliferation of cells around taste buds in a protective manner during post-irradiation recovery. Mechanically, we revealed that the most frequent m6A modifications were enriched in Hippo and Wnt signaling, and specific peaks were observed near the stop codons of Lats1 and FZD7. Our study elucidates that METTL3 is essential for taste bud formation and could promote the quantity recovery of taste bud after radiation.

OShnscc: a novel user-friendly online survival analysis tool for head and neck squamous cell carcinoma based on RNA expression profiles and long-term survival information / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Head and neck squamous cell carcinoma (HNSCC), as the most common type (>90%) of head and neck cancer, includes various epithelial malignancies that arise in the nasal cavity, oral cavity, pharynx, and larynx. In 2020, approximately 878 ‍ 000 new cases and 444 000 deaths linked to HNSCC occurred worldwide (Sung et al., 2021). Due to the associated frequent recurrence and metastasis, HNSCC patients have poor prognosis with a five-year survival rate of 40%-50% (Jou and Hess, 2017). Therefore, novel prognostic biomarkers need to be developed to identify high-risk HNSCC patients and improve their disease outcomes.