Abstract:This review provides a comprehensive summary of the latest advancements in high-throughput protein structural bioinformatics, a field that has undergone a revolutionary transformation with the advent of deep learning-based protein structure prediction systems like AlphaFold2. These systems have significantly increased the accuracy, speed, and scale of protein structure prediction, resulting in an exponential growth in the number of protein structures available for analysis. Notably, the AlphaFold Protein Structure Database (AFDB) has amassed over 214 million protein structures, surpassing the PDB’s 50-year cumulative data by over 1 000-fold within several months. Big data is driving the comprehensive upgrade of protein structural bioinformatics. This review focuses on three main areas: structure data management, tool development, and structure data mining. In the realm of structure data management, the review spotlights the optimization strategy of AlphaFold-like systems, which significantly reduces the resource requirements for protein folding, enabling more researchers to make custom structure predictions and further enlarging the data scale. The resulting “data explosion” has exerted increased pressure on storage and bandwidth, prompting the development of cutting-edge tools such as Foldcomp, PDC, and ProteStAr for compressing PDB files. Moreover, the review underscores the critical role of public repositories like ModelArchive and PDB-Dev in archiving and sharing third-party AlphaFold models. It also highlights the utilization of independent services like MineProt and 3D-Beacons to create more interactive and accessible data portals. In terms of tool development, the review spotlights recent breakthroughs in structure alignment algorithms, represented by Foldseek, which enable ultra-fast searching of large protein structure databases. It also covers tools for functional annotation of proteins based on their structures, including AlphaFill for ligand annotation, DeepFRI for Gene Ontology (GO) annotation, TT3D for protein-protein interaction (PPI) prediction, among others. It is proposed that 3Di sequences born concurrently with Foldseek can enhance many sequence-based deep learning models developed in the pre-AlphaFold era, enabling them to be applied to structure-based function prediction. The challenges on traditional molecular docking methods in the high-throughput era are mentioned at last, in a gesture to arouse the attention of researchers. Finally, the review explores the burgeoning field of structure data mining. Whole proteome structuring has become feasible in recent years, and scientists are processing large structure datasets from an omics viewpoint, continuously identifying analyzable elements and optimizing methodologies, as well as utilizing newly developed tools to push the boundaries. Notable examples include the identification of new protein families, the development of protein structure clustering, and the integration of AlphaFold with conventional experimental techniques to solve large structures. These advancements are paving the way for a deeper understanding of protein structure and function and have the potential to unlock new discoveries in the life sciences. However, the review also acknowledges the challenges and limitations that persist in the field, including the lack of diversity in high-throughput software for protein structural bioinformatics and the existing bottleneck in rapidly predicting protein complex structures. Overall, structural bioinformatics is expected to play an even more crucial role in the life sciences with the development of high-throughput methodology.

Abstract:Given the constraints imposed by the “14-day ethics” rule, numerous critical events occurring between the second and fourth weeks of embryonic development remain poorly understood. This underscores the necessity of a detailed understanding of embryonic development and regulation during this period, which is indispensable for preventing pregnancy failure, treating birth defects, and promoting human reproductive health.Rodents, characterized by their small size, rapid growth, strong reproductive capacity, and fully sequenced genomes, are widely used as crucial models for studying embryonic development. However, the substantial physiological differences between rodents and primates due to evolutionary divergence make it challenging to directly apply findings from rodent studies to primates. Besides, primates, our closest relatives in terms of evolutionary phylogenetics and physiological characteristics, share more than 95% genetic homology with humans, underscoring the urgent need for primate research. Furthermore, early-stage embryonic cells are both scarce and diverse, making their regulatory mechanisms and developmental pathways typically elucidated through single-cell sequencing. For instance, three significant articles published in Science in 2018 mapped the complete atlas of organ and tissue development from fertilization and captured dynamic gene expression profiles in zebrafish and frogs through single-cell transcriptomics. Unfortunately, relying solely on single-cell omics analysis falls short in effectively and comprehensively deciphering the intricate cellular network information. Single-cell multi-omics empower researchers to systematically decode cell heterogeneity and developmental trajectories at the individual cell level by combining transcriptomics, epigenomics, proteomics, and metabolomics analyses. These emerging technologies play a significant role in life sciences, enabling the elucidation of critical early primate embryonic development events from a multi-dimensional perspective, including zygotic genome activation (ZGA), X-chromosome dosage compensation, origins of primordial germ cells (PGCs), mechanisms of cell fate determination, and pivotal events in gastrulation and early organogenesis.This article chronicles the advancement of pivotal technologies, from single-cell histology to multi-omics, beginning with the single-cell transcriptome and culminating in a comprehensive analysis according to the central dogma of molecular biology. It highlights the transition from a singular to a holistic perspective in cellular analysis and reviews the application of multi-omics techniques in unveiling early primate embryonic development. Finally, it delves into the application of multi-omics technologies in enhancing our understanding of early primate embryonic development and explores future possibilities, directions, and challenges in this rapidly evolving field. In doing so, it emphasizes the critical role of interdisciplinary approaches, combining insights from genetics, molecular biology, and bioinformatics to foster innovations in reproductive medicine and developmental biology. The integration of such technologies offers the promise of breakthroughs in understanding complex biological processes, potentially leading to novel therapeutic strategies and advancements in reproductive health and medicine.

Abstract:Consciousness and unconsciousness represent a compelling topic in psychology and neuroscience, embodying a dynamic interplay between two fundamental cognitive states. Understanding the cognitive and neural mechanisms underlying their interaction poses a significant challenge. While previous studies have predominantly focused on the impact of attention on consciousness, the influence of attention on unconscious processes has often been overlooked. However, the role of attention in modulating unconscious information processing is paramount, as it can regulate various aspects of unconscious visual processing, including simple visual information, semantic content, and emotional stimuli. Within the visual processing pathway, attention operates at early levels to modulate unconscious visual processing, starting at least from the eye-of-origin and visual orientations. In the semantic system, attention can top-down enhance unconscious semantic processes in a goal-dependent manner, enhancing goal-relevant processes while suppressing goal-irrelevant ones. In the emotional system, attentional load, in addition to target relevance, can regulate unconscious emotional processing. These findings suggest that the regulatory role of attention on unconscious processes depends on both goal relevance and the amount of attentional resources. Specifically, the goal-relevance of unconscious processes determines the direction of attentional modulation, while the amount of attentional resources allocated determines the extent of modulation. The once-prevailing notion that unconscious processing is automatic and not subject to attentional modulation has been gradually overturned. Current studies indicate that attention can modulate both conscious and unconscious processes, providing a new perspective on the relationship between attention and consciousness. Spatial attention can operate independently from consciousness at the neural representation level. Furthermore, other factors tightly related to attention, such as goal-related task sets, working memory, and attentional load, can all impact unconscious processes. These findings collectively suggest that attention and consciousness are functionally dissociated, supporting the idea that attention is necessary for both conscious and some unconscious processes. In conclusion, unconscious information processing is a complex and intriguing field where attention plays a crucial role. Continued in-depth research in this area is needed to deepen our understanding of how the human brain processes unconscious information and how attention exerts its regulatory influence. This not only requires studying the commonalities and specificities of different types of attention but also examining the sharing and individuality among different sensory modalities and cognitive modules. Theoretically, this not only helps us understand the mechanisms of attention but also sheds light on the mechanisms of consciousness. Studying these issues is also of practical value. Importantly, the organization and regulation of unconscious processes are closely related to human survival and development. For example, while rapid unconscious emotional processes (such as unconscious fear) are beneficial for rapid threat responses and increased survival chances, excessive and uncontrolled unconscious emotional processes can lead to anxiety disorders, phobias, and other mental disorders. Furthermore, while repeated perceptual and behavioral training can improve efficiency by forming highly automated unconscious processes, excessively stubborn unconscious processes can hinder the learning of new skills. Studying the role of attention in regulating these unconscious processes can help develop new intervention methods to maintain mental health and improve behavioral performance.

Abstract:As polar cells, neurons are composed of a cell body, dendritic networks, and long, branched axons. To maintain normal physiological functions throughout the lifespan of vertebrates, differentiated neurons require substantial energy to sustain resting potential and synaptic transmission. Neurons predominantly rely on ATP generated through mitochondrial oxidative phosphorylation for energy. They transport and accumulate healthy mitochondria to energy-demanding areas, such as the presynaptic terminals of axon branches, through long-distance transport and anchoring, while reversing the transport of aged or damaged mitochondria in the axon terminals back to the soma for degradation. This article, integrating authors’ research, discusses from a mechanical perspective how mitochondria overcome resistance to achieve long-distance transport along axons under the influence of driving forces. The review covers topics such as microtubule polarity, microtubule motor proteins, mitochondrial docking protein complexes, interactions between mitochondria and anchoring proteins, intracellular resistance, interactions between mitochondria and the endoplasmic reticulum, and aspects of mitochondrial biogenesis, fission, fusion, division, and quality control. These novel perspectives will provide important insights for understanding neurological diseases caused by mitochondrial transport dysfunctions.

Abstract:Macrophages, existed in almost all organs of the body, are responsible for detecting tissue injury, pathogens, playing a key role in host defense against a variety of invading pathogens triggering inflammatory responses, and emerging evidence suggests that macrophage-mediated immune responses are efficiently regulated by the ubiquitination modification, which is responsible for normal immune responses. However, numerous studies indicates that the aberrant activation or inhibition of macrophage-mediated immune responses occurs in inflammation, mainly caused by dysregulated ubiquitination modification due to E3 ubiquitin ligases mutations or abnormal expression. Notably, E3 ubiquitin ligases, responsible for recognizing the substrates, are key enzymes in the ubiquitin-proteasome system (UPS) composed of ubiquitin (Ub), ubiquitin-activating E1 enzymes, ubiquitin-conjugating E2 enzymes, E3 ubiquitin ligases, 26S proteasome, and deubiquitinating enzymes. Intriguingly, several E3 ubiquitin ligases are involved in the regulation of some common signal pathways in macrophage-mediated inflammation, including Toll-like receptors (TLRs), nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs), RIG-I-like receptors (RLRs), C-type lectin receptors (CLRs) and the receptor for advanced glycation end products (RAGE). Herein, we summarized the physiological and pathological roles of E3 ligases in macrophage-mediated inflammation, as well as the inhibitors and agonists targeting E3 ligases in macrophage-mediated inflammation, providing the new ideas for targeted therapies in macrophage-mediated inflammation caused aberrant function of E3 ligases.

Abstract:Hepatocellular carcinoma is one of the most common malignant tumors worldwide, posing a great threat to human health and life. Despite the tremendous progress in understanding the origin and molecular characterization of hepatocellular carcinoma, there are still few therapeutic options that can significantly increase the survival rate and improve the quality of life of patients. Protein post-translational modifications (PTMs) are regulatory mechanisms for protein activity, localization, expression, and interactions with other cellular molecules that induce changes in protein properties and functions. More and more studies have demonstrated that PTMs and immunotherapy play an important role in the development of hepatocellular carcinoma, even in the immunosurveillance of hepatocellular carcinoma and the treatment and prognosis of hepatocellular carcinoma patients. Traditional types of PTMs include phosphorylation, glycosylation, methylation, and ubiquitination. Phosphorylation affects cancer development and progression by regulating tumor cell proliferation, invasion and metastasis, and inhibiting apoptosis. There are two main types of glycosylation: O-glycosylation and N-glycosylation. Abnormal glycosylation not only promotes the proliferation and metastasis of hepatocellular carcinoma cells, but also plays an important role in immune recognition and immune escape. Common methylation modifications include DNA methylation, RNA methylation and histone methylation. Among them, histone methylation, as an important epigenetic regulatory mechanism, is of great theoretical and practical significance for understanding the mechanism of hepatocellular carcinoma as well as carrying out the corresponding prevention and immunotherapy. Ubiquitination plays an important role in the localization, metabolism, function, regulation and degradation of proteins, and it is regulated at different levels by ubiquitin-activating enzyme (E1), ubiquitin-conjugating enzyme (E2), ubiquitin-conjugating enzyme (E3), and a series of deubiquitinating enzymes (DUBs) and is closely related to hepatocellular carcinoma immunotherapy. This paper begins with a brief overview of the importance of PTMs of proteins, discusses the importance of these traditional types of PTMs in hepatocellular carcinoma immunotherapy, and summarizes the most recent applications of these approaches in hepatocellular carcinoma in order to explore the mechanism of action of PTMs in hepatocellular carcinoma immunotherapy. Then, we summarize the finding that programmed death-ligand 1 (PD-L1) is associated with a variety of conventional types of PTMs, that in-depth study of the mechanisms regulating PD-L1 expression in tumor cells is expected to improve therapeutic efficacy, and that targeting PD-L1 in PTMs is expected to be a new field for exploring hepatocellular carcinoma immunotherapy in the future. Finally, we discuss the current status of research on PTMs for hepatocellular carcinoma immunotherapy and provide new insights and future research directions. In addition to the traditional types of PTMs, multiple novel PTMs have also been identified in published research reports, while the relationship between novel PTMs and hepatocellular carcinoma and the types of PTMs to other undiscovered proteins are still poorly understood, and future research will be focused on a more comprehensive knowledge and understanding of PTMs as well as on exploring new types and mechanisms of PTMs. Overall, further investigation of the role of PTMs in tumor immunity could help to discover new biomarkers and to develop more effective and personalized cancer immunotherapies and targeted therapies, expanding our understanding of cancer biology.

Abstract:Cardiovascular and cerebrovascular diseases, usually result from atherosclerosis, has the highest mortality rate globally. Lipid metabolism disorder is the main cause of atherosclerotic cardiovascular and cerebrovascular diseases, which not only lead to acute diseases such as myocardial infarction, stroke, acute pancreatitis, but also chronic kidney disease. In recent years, the advancement of gene therapy technologies has provided novel means for lipid metabolism study, and has also made it possible to cure patients with congenital lipid metabolism abnormalities. Adeno-associatd virus has a wide host range, high safety, low immunogenicity, and especially the ability of long-term stable expression in vivo, making it the preferred delivery tool for gene therapy of monogenic genetic diseases. Alipogene triprivec, also known as Glybera, was approved by the European Medicines Agency in 2012. It is the first gene therapy drug that uses recombinant AAV1 vector to directly deliver a highly active LPL protein S447X mutant to muscle cells for the treatment of patients with hereditary LPL deficiency. To enhance the targeted transduction efficiency of AAV carriers, recombinant AAV8.TBG.hLDLR utilizes the tissue tropsim of AAV8 to liver, meanwhile utilizes a liver specific thyroxine binding globulin promoter to control gene transcription, thereby achieving liver cell specific high expression of human low-density lipoprotein receptors (LDLR). In patients with familial hypercholesterolemia, AAV8.TBG.hLDLR treatment effectively lower the level of plasma LDL for a long time, thus preventing the occurrence of atherosclerosis.Proprotein convert subunit kexin 9 (PCSK9) is secreted by liver cells. PCSK9 binds and transports LDLR to lysosomes for degradation, preventing the circulation and regeneration of LDLR, leading to accelerated degradation of LDLR and finally resulting in the accumulation of low-density lipoprotein cholesterol in plasma.Using AAV to deliver Cas9 of Staphylococcus aureus and gRNA targeting the Pcsk9 gene can knock out Pcsk9 in mouse liver, leading to a long-term significant decrease in plasma cholesterol levels in mice. Hepatocyte specific angiopoietin related protein 3 (Angptl3) is an endogenous inhibitor of LPL. Using the AAV9 mediated AncBE4max system and the dCas9 mediated single base gene editing system to introduce early termination codons, the knockout of Angptal3 in liver cells was achieved with an average knockout efficiency of 63.3%. After 2-4 weeks of administration in mice, the Angptl3 protein was completely undetectable in the peripheral blood, and serum triglycerides and total cholesterol decreased by 58% and 61%, respectively. Ring finger containing protein 130 (RNF130) is an E3 ubiquitin ligase. Research has shown that overexpression of RNF130 using AAV2/8 leads to ubiquitination degradation and redistribution of LDLR on the cell membrane, significantly reducing LDLR expression on liver cells and increasing plasma LDLC levels, while knocking out Rnf130 gene using the AAV-CRISPR system results in the opposite effect. This AAV mediated RNF130 function study proves that RNF130 is a posttranslational regulatory protein of LDLR and plays an important role in the regulation of serum LDLC. As mentioned above, recently, various lipid-lowering gene therapy drugs carried by different serotypes of adeno-associated virus have been applied in clinic or are undergoing clinical trials, and adeno-associated virus has emerging to be an important tool for lipid metabolism research.This article reviews the new progress of adeno-associated virus vectors in lipid metabolism study and lipid-lowering gene therapy.

Abstract:Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease, defined by several phases, ranging from benign fat accumulation to non-alcoholic steatohepatitis (NASH), which can lead to liver cancer and cirrhosis. Although NAFLD is a disease of disordered metabolism, it also involves several immune cell-mediated inflammatory processes, either promoting and/or suppressing hepatocyte inflammation through the secretion of pro-inflammatory and/or anti-inflammatory factors to influence the NAFLD process. However, the underlying disease mechanism and the role of immune cells in NAFLD are still under investigation, leaving many open-ended questions. In this review, we presented the recent concepts about the interplay of immune cells in the onset and pathogenesis of NAFLD. We also highlighted the specific non-immune cells exhibiting immunological properties of therapeutic significance in NAFLD. We hope that this review will help guide the development of future NAFLD therapeutics.

Abstract:Pancreatic cancer (PC) is a highly fatal disease which originated from pancreatic epithelial and acinar cells, and the survival rate of pancreatic cancer patients is only about 12%. Approximately 95% of pancreatic cancer presents as ductal adenocarcinoma (PDAC). Pancreatic cancer is characterized by high aggressiveness, rapid progression and progression, and high resistance to treatment. Common somatic mutated genes in the early stage of pancreatic cancer include KRAS, CDKN2A, TP53, and SMAD4. Most pancreatic cancer patients are affected by environmental risk factors such as age, sex and diet. Malignant pancreatic cancer is associated with non-invasive, preneoplastic lesions that are thoughted to be precursors, such as pancreatic intraepithelial neoplasia (PanIN), intraductal papillary mucinous neoplasm (IPMN) and mucinous cystadenoma (MCN). In recent years, people have gradually improved the therapy and diagnosis of pancreatic cancer, and the contribution of imaging technology, which enhancing the usage of minimally invasive pancreatectomy that typically includes pancreaticoduodenectomy and distal pancreatectomy. However, combined administration of the chemotherapeutic gemcitabine and erlotinib is still considered a potential first-line treatment for advanced pancreatic cancer, but the development of chemoresistance often leads to poor therapeutic outcomes. Based on the current research progress for pancreatic cancer, its treatment currently remains one of the most important challenges in the medical field. Although some new treatment options have been provided, there were minor clinical success achieved and therefore new safe and effective therapies of pancreatic cancer are still an urgent need for patients. Among these new therapies for pancreatic cancer, short peptide-based treatment protocols have attracted great attention. Peptide is a compound formed by linking α-amino acids together in peptide chains. It is also an intermediate product of proteolysis. The short peptide-based therapy has many advantages such as precise targeting, easy preparation and low toxicity. Short peptides usually act as tumor suppressors by targeting and recognizing tumor-specific expressed proteins. Currently, there is an increased interest in peptides in pharmaceutical and development research, and approximate 140 peptide therapeutics are currently being evaluated in clinical trials. These peptides provide excellent prospects for targeted drug delivery because of their high selectivity, specificity and simplicity of modification. Peptides have high bioactivity and excellent biodegradability. Clinically, short peptides are increasingly used as combination drugs with chemotherapy for tumor treatment. Peptides can induce cancer cell death by numerous mechanisms and peptides have emerged as a promising drug for the treatment of pancreatic cancer. Here we mainly review the roles of peptides on Wnt/β-catenin, NF-κB, autophagy, and the use of peptides as tracer in pancreatic cancer. We also analyzed the benefits and disadvantages existing in the development process of short peptides, which provide the feasibility of targeted short peptides to become new therapeutic approaches for cancer therapy.

Abstract:mRNA vaccines and drugs enter host cells through delivery vectors and produce target proteins using the protein synthesis mechanism of cells. mRNA and target proteins can induce the body to produce innate immunity and adaptive immunity, and the target protein itself can also play a corresponding role. Tumor cells are inhibited and cleared under the above immune effects and target proteins. This article reviews the immunogenicity of mRNA, that is, the specific mechanism of stimulating the body to produce an immune response.At the same time, the main types of cells transfected by mRNA vaccine were briefly introduced. (1) Muscle cells, epidermal cells, dendritic cells and macrophages at the injection site; (2) immune cells in peripheral lymphoid organs; (3) liver cells and spleen cells, etc. Although transfected with a variety of cells, it is mainly enriched in immune cells and liver cells because immune cells express toll-like receptors and liver cells express low-density lipoprotein receptors. mRNA vaccines and drugs are mainly divided into non-replicating mRNA (nrmRNA), self-amplifying RNA (saRNA), trans-amplifying RNA (taRNA) and circular RNA (circRNA).This article reviews how these 4 types of vaccines and drugs work, and compares their advantages and disadvantages. Due to its inherent immunogenicity, instability, and low delivery efficiency in vivo, mRNA vaccines and drugs have been unable to enter the clinic. This article describes in detail how to reformation and modify the 5"cap, 5"UTR, 3"UTR, ORF, 3"Poly(A) and some nucleotides of mRNA to eliminate its immunogenicity and instability. Due to the low efficiency of the delivery carrier, the researchers optimized it. This article briefly introduces the application of non-viral vectors and their targeting, specifically involving the mechanism of action of various types of delivery vectors and their advantages and disadvantages, and summarizes some of the current targeting vectors. Targeted carriers can improve the delivery efficiency of mRNA to specific tissues and prevent side effects of systemic exposure, such as liver injury. The specific methods of using mRNA vaccines and drugs to treat cancer are as follows: mRNA can be used to encode and transcribe tumor-associated antigens, tumor-specific antigens (TSAs), therapeutic antibodies, cytokines, tumor suppressors, oncolytic viruses, CRISPR-Cas9, CARs and TCRs, so as to play an anti-tumor role. In this paper, the specific mechanism of the above methods and the current research and development of corresponding mRNA vaccines and drugs are briefly reviewed. The successful development of the COVID-19 mRNA vaccine has brought mRNA technology to the attention of the world and brought new and effective means for the prevention and treatment of cancer. mRNA vaccines and drugs have the advantages of short development cycle, dual immune mechanism, safety, high efficiency and large-scale production. At the same time, there are also many areas that need further improvement, such as the development of ideal target TSAs, the in-depth development of saRNA, taRNA and circRNA, the development of targeted nano-delivery for different tissues and organs, the expansion of mRNA administration routes, and the development of mRNA that can be stably stored at room temperature or even high temperature. These problems need to be further studied and solved. In addition to cancer therapy, mRNA vaccines and drugs can also be used in the treatment of infectious diseases, genetic diseases, regenerative medicine and anti-aging. mRNA vaccines and drugs are a very promising platform, and we believe that they will benefit cancer patients in the near future.

Abstract:The relationship between exercise and cardiac health has always been a hotspot in the fields of medicine and exercise science. Recently, with the in-depth study of the biological clock, people have gradually realized the close relationship between cardiac metabolic activity and circadian rhythms. The mammalian circadian system includes the central circadian clock and peripheral circadian clocks, the central circadian clock is the main clock system responsible for regulating the circadian rhythms in organisms, located in the suprachiasmatic nucleus (SCN) of the hypothalamus in mammals, which receives light signals from the retina and translates them into neural signals to regulate peripheral circadian clocks distributed throughout the body. Peripheral circadian clocks exist in various tissues and organs of organisms, coordinating with the central circadian clock to maintain the circadian rhythms of the organism. A series of clock genes regulate downstream clock-controlled genes through the transcriptional-translational feedback loop (TTFL), profoundly affecting the physiological activities of the heart, including cardiac contraction, relaxation, and metabolic processes. Factors such as sleep disorders, shift work, light pollution, and excessive use of electronic devices in modern lifestyles have led to widespread disruption of circadian rhythms, which are significantly correlated with increased cardiovascular disease incidence and mortality. Studies have found that dysregulation of the cardiac circadian clock can not only lead to myocardial lipid degeneration and weakened metabolic rhythms but also decrease myocardial glucose utilization, thereby increasing the risk of adverse cardiac events. Exercise, as a key zeitgeber, has been widely demonstrated to regulate the circadian clocks of peripheral organs such as skeletal muscle, kidneys, and liver. Additionally, exercise, as an important means to improve cardiovascular function, can effectively enhance cardiac metabolic function and resistance to stress stimuli, playing a significant role in promoting heart health. However, the specific mechanisms by which exercise affects the cardiac circadian clock and its related genes are currently unclear. Therefore, this review will focus on the relationship between the cardiac circadian clock and cardiac metabolic activity, summarize previous research to review the possible mechanisms of exercise-mediated regulation of cardiac metabolic activity on the cardiac circadian clock. The cardiac circadian clock plays an important role in maintaining cardiac metabolic activity and physiological functions. The loss of cardiac circadian clock genes Bmal1 and Clock can significantly reduce cardiac fatty acid and glucose utilization rates, increase myocardial lipotoxicity, weaken the circadian rhythm of myocardial triglyceride metabolism, and lead to abnormalities in the circadian clocks of other peripheral organs. Exercise, as a zeitgeber, can independently regulate the cardiac circadian clock apart from the central circadian clock. Additionally, exercise, as an important means to improve cardiovascular function, may regulate cardiac metabolic activity and the transcription of clock genes by activating the hypothalamic-pituitary-adrenal axis (HPA) and sympathetic-adrenal-medullary axis (SAM) and regulating energy metabolism, thereby maintaining the stability of the cardiac circadian clock and promoting heart health. Future research on the molecular mechanisms of exercise regulation of the cardiac circadian clock will help clarify the role and impact of clock genes in cardiac metabolism and physiological activities, providing new preventive and treatment strategies for shift workers, night owls, and patients with cardiovascular diseases. Therefore, future research should focus on (1) the mechanisms by which exercise regulates cardiac metabolic activity and the circadian clock, (2) the effects and mechanisms of exercise on the disruption of cardiac circadian clock induced by light-dark cycle disturbances, and (3) the effects of exercise on the metabolic activity and circadian rhythms of other peripheral organs regulated by the cardiac circadian clock.

Abstract:Smoking is the leading preventable risk factor for disease and death worldwide. Tobacco and its smoke contain a complex mix of over 9 500 chemical substances, including oxidative gases, heavy metals, and 83 known carcinogens. Long-term smoking is a significant risk factor for respiratory diseases such as acute lung injury, emphysema, and pulmonary fibrosis. Damage to alveolar epithelial cells (AECs) is a common pathological feature in these smoking-related lung diseases. AECs, which line the surface of the alveoli, play a crucial role in preventing overexpansion or collapse, secreting cell factors and surfactants, containing abundant mitochondria, and being essential for lung tissue maturation, gas exchange, metabolism, and repair after damage. Damage to these cells can lead to pulmonary edema and alveolar collapse. Cigarette smoke (CS) can disrupt alveolar epithelial cell function through various pathways, resulting in cell death, tissue damage, and the development of lung diseases.This review summarizes recent research on the damage caused by CS to AECs, showing that CS can promote cell death and damage through induction of oxidative stress, autophagy, endoplasmic reticulum stress, mitochondrial dysfunction, inflammation, and epithelial-mesenchymal transition. It also affects the proliferative function of alveolar type II epithelial cells. The review highlights that CS-induced oxidative stress is a key factor in causing various types of damage, with TRP ion channels serving as important triggers. Inhibiting CS-induced oxidative damage can significantly prevent cell death and subsequent diseases such as pulmonary emphysema. The activation of the same pathway induced by CS can lead to different types of cell damage, potentially encouraging the development of different diseases. CS can either directly induce or indirectly promote cell inflammation through endoplasmic reticulum stress, mitochondrial dysfunction, and senescence. There are interconnected relationships between these mechanisms, and SIRT1 is an important protein in preventing CS-induced AECs damage. Increasing SIRT1 activity can alleviate CS-induced autophagy, endoplasmic reticulum stress, and senescence in various cell damages; its substrate NAD⁺ is already used clinically, and its effectiveness in COPD treatment deserves further exploration. The impact of CS on cells varies based on concentration: lower concentrations stimulate stress responses or apoptosis, while higher concentrations lead to apoptosis or necrosis through various mechanisms, ultimately impairing lung epithelial function. When external stimuli exceed the cells’ self-healing capacity, they can cause damage to cells, lung epithelial barriers, and alveoli, promoting the development of related lung diseases. Key proteins that play a protective role may serve as potential targets to mitigate cell damage.This review provides insights into the various mechanisms through which CS induces damage to AECs, covering important transcription factors, DNA repair proteins, and membrane channel proteins, paving the way for the study of new mechanisms and pathways. However, there are still unanswered questions, such as the need for further exploration of the upstream pathways of CS-induced autophagy in AECs and the intrinsic mechanisms of CS in enhancing the stem cell properties of AECs and its relationship to the occurrence of lung cancer.It is expected that this article will provide a theoretical basis for future research on the mechanisms of lung epithelial cell damage caused by CS or its individual components and inspire clinical strategies for the prevention and treatment of smoking-related lung diseases.

Abstract:DNA genetic markers have always played important roles in individual identification, kinship analysis, ancestry inference and phenotype characterization in the field of forensic medicine. DNA methylation has unique advantages in biological age inference, body fluid identification and prediction of phenotypes. The majority of current studies independently examine DNA and DNA methylation markers using various workflows, and they use various analytical procedures to interpret the biological information these two markers present. Integrated methods detect DNA and DNA methylation markers simultaneously through a single experimental workflow using the same preparation of sample. Therefore, they can effectively reduce consumption of time and cost, streamline experimental procedures, and preserve valuable DNA samples taken from crime scenes. In this paper, the integrated detection approaches of DNA and DNA methylation markers on different detection platforms were reviewed. In order to convert methylation modifications to detectable forms, several options were available for pretreatment of genomic DNA, including digestion with methylation-sensitive restriction enzyme, affinity enrichment of methylated fragments, conversion of methylated or unmethylated cytosine. Multiplexed primers can be designed for DNA markers and converted DNA methylation markers for co-amplification. The schemes of using capillary electrophoresis platform for integrated detection add the pretreatment of genomic DNA on the basis of detecting DNA genetic markers. DNA and DNA methylation markers are then integrated by co-amplification. But the limited number of fluorescent options available and the length of amplicons restrict the type and quantity of markers that can be integrated into a panel. Pyrophosphate sequencing also supports integrated detection of DNA and DNA methylation markers. On this platform, due to the conversion of unmethylated cytosine to thymine after treatment with bisulfite, the methylation level of CpG site can be directly calculated using the peak height ratio of cytosine bases and thymine bases. Therefore, the methylation levels and SNP typing can be simultaneously obtained. However, due to the limited read length of sequencing, the detection of markers with longer amplicons is restricted. It is not conducive to fully interpret the complete information of the target sequence. Next-generation sequencing also supports integrated detection of DNA and DNA methylation markers. A preliminary experimental process including DNA extraction, pretreatment of genomic DNA, co-preparation of DNA and DNA methylation library and co-sequencing, has been formed based on the next-generation sequencing platform. It confirmed the feasibility of next-generation sequencing technology for integrated detection of DNA and DNA methylation markers. In field of biomedicine, various integrated detection schemes and corresponding data analysis approaches of DNA and DNA genetic markers developed based on the above detection process. Co-analysis can simultaneously obtain the genomic genetic and epigenetic information through a single analytic process. These schemes suggest that next-generation sequencing may be an effective method for achieving more accurate and highly integrated detection, helping to explore the potential for application in forensic biological samples. We finally explore the impact of interactions between sites and different pretreatment methods on the integrated detection of DNA and DNA methylation markers, and also propose the challenge of applying third-generation sequencing for integrated detection in forensic samples.

Abstract:Objective To investigate the effect of 8-week aerobic exercise on the improvement of core behaviors of male and female autistic mice induced by valproic acid (VPA).Methods Experimental mice were randomly divided into the control group (CTL), VPA-induced autism group (VPA) and VPA+aerobic exercise group (VEX), with 10 mice in each group. The pregnant mice were injected with VPA intraperitoneally at E12.5, and their offspring were used as autistic mice. Pups were weaned 28 d after birth and began an 8-week aerobic exercise intervention. The day after exercise, mice were tested in behavioral experiments to detect exploratory behavior, social skills, repetitive stereotypic behavior, cognitive ability and mood. The mice were tested for social skills, repetitive stereotyped behaviors, cognitive and learning memory abilities, exploratory behaviors, and emotions by behavioral assays on the following day after the exercise.Results Both male and female mice in the CTL group showed a significant decrease in the total distance and percentage of time spent in the interaction zone in the 4th socialization compared to the 1st socialization (P<0.01); the total distance and percentage of time spent in the interaction zone in the 5th socialization was significantly increased compared to the 4th socialization (P<0.01); in VPA group, both male and female mice showed no significant change in the total distance and percentage of time spent in the interaction zone in the 4th and 5th socialization; in the VEX group, the total distance and percentage of time spent in the interaction zone by male mice in the 4th socialization was significantly decreased compared to the 1st socialization (P<0.01, P<0.05); and in the VEX group the total distance and percentage of time spent in the social interaction zone by both male and female mice in the 5th socialization was significantly increased compared to the 4th socialization (P<0.01, P<0.05). The results of the first phase of three-box socialization experiment showed that male and female mice in the CTL group spent more time socializing with their social partners than in contact with the empty cages (P<0.01); there was no difference in the time spent by male and female mice in the VPA group in socializing with their social partners and the empty cages; and male and female mice in the VEX group spent a longer time socializing with their social partners (P<0.01). The results of the second phase of three-box test showed that male and female mice in the CTL group showed a significant tendency to socialize with new social partners (P<0.01), whereas no significant changes were observed in the mice of VPA group; aerobic exercise significantly ameliorated this deficit in male and female mice with autism. Compared with the CTL group, VPA-induced significant decreases were observed in the total distance freely moved in the central area of the open field, the time and percentage of time spent in the open arm of cross maze, and the distance and time spent in the white box in both male and female autistic mice (P<0.01); a significant increase in the number of bead burials and time spent in self-grooming (P<0.01); a significant decrease in the cognitive index (P<0.01); a significantly longer latency to find the platform, and significantly decreased the percentage of time spent in the target quadrant and the number of times they traversed the platform (P<0.01). Compared with the VPA group, after 8 weeks of aerobic intervention, male and female mice in the VEX group showed a significant increase in total distance, open-arm dwell time, and percentage of free movement in the central area of the empty field (P<0. 05), and a trend toward a decrease in the dwell time of females in the white box was not significant, the number of beads burying and the time of self-combing were significantly lower (P<0.01, P<0.05); and a significant increase in cognitive index (P<0.05), a significantly shorter time to find the platform, and significantly increased percentage of time spent in the target quadrant and the number of times they traversed the platform (P<0.01), showing excellent learning memory.Conclusion Autistic mice severely suffer from social and cognitive impairments, repetitive stereotyped behaviors, decreased activity level, and the exhibition of anxiety. 8 weeks of aerobic exercise can improve the social and cognitive abilities, alleviate the stereotyped repetitive behaviors, increase the activity level, and positively regulate the anxiety in autistic mice. It is hypothesized that aerobic exercise has an important role in motor rehabilitation of autism, in order to provide a theoretical basis for clinical research.

Abstract:Objective To investigate the effect of mucin 1 (MUC1) on the proliferation and apoptosis of nasopharyngeal carcinoma (NPC) and its regulatory mechanism.Methods The 60 NPC and paired para-cancer normal tissues were collected from October 2020 to July 2021 in Quanzhou First Hospital. The expression of MUC1 was measured by real-time quantitative PCR (qPCR) in the patients with PNC. The 5-8F and HNE1 cells were transfected with siRNA control (si-control) or siRNA targeting MUC1 (si-MUC1). Cell proliferation was analyzed by cell counting kit-8 and colony formation assay, and apoptosis was analyzed by flow cytometry analysis in the 5-8F and HNE1 cells. The qPCR and ELISA were executed to analyze the levels of TNF-α and IL-6. Western blot was performed to measure the expression of MUC1, NF-кB and apoptosis-related proteins (Bax and Bcl-2).Results The expression of MUC1 was up-regulated in the NPC tissues, and NPC patients with the high MUC1 expression were inclined to EBV infection, growth and metastasis of NPC. Loss of MUC1 restrained malignant features, including the proliferation and apoptosis, downregulated the expression of p-IкB、p-P65 and Bcl-2 and upregulated the expression of Bax in the NPC cells.Conclusion Downregulation of MUC1 restrained biological characteristics of malignancy, including cell proliferation and apoptosis, by inactivating NF-κB signaling pathway in NPC.

Abstract:Objective The absorption of substances into blood is mainly dependent on the mesenteric lymphatic pathway and the portal venous pathway. The substances transported via the portal venous pathway can be metabolized by the biotransformation in the liver. On the contrary, the substances in the mesenteric lymph fluid enter the blood circulation without biotransformation and can affect the body directly. Alcohol consumption is strongly linked to global health risk. Previous reports have analyzed the changes of metabolites in plasma, serum, urine, liver and feces after alcohol consumption. Whether alcohol consumption affects the metabolites in lymph fluid is still unknown. Therefore, it is particularly important to explore the changes of substances transported via the mesenteric lymphatic pathway and analyze their harmfulness after alcohol drinking.Methods In this study, male Wistar rats were divided into high, medium, and low-dosage alcohol groups (receiving Chinese Baijiu at 56%, 28% and 5.6% ABV, respectively) and water groups. The experiment was conducted by alcohol gavage lasting 10 d, 10 ml·kg^-1·d^-1. Then mesenteric lymph fluid was collected for non-targeted metabolomic analysis by using liquid chromatography-mass spectrometry (LC-MS) and bioinformatic analysis. Principal component analysis and hierarchical clustering were performed by using Biodeep. Meanwhile, KEGG enrichment analysis of the differential metabolites was also performed by Biodeep. MetaboAnalyst was used to analyze the relationship between the differential metabolites and diseases.Results The metabolites in the mesenteric lymph fluid of the high-dosage alcohol group change the most. Based on the KEGG enrichment analysis, the pathways of differential metabolites between the high-dosage alcohol group and the control group are mainly enriched in the central carbon metabolism in cancer, bile secretion, linoleic acid metabolism, biosynthesis of unsaturated fatty acids, etc. Interestingly, in the biosynthesis of unsaturated fatty acids category, the content of arachidonic acid is increased by 7.25 times, whereas the contents of palmitic acid, oleic acid, stearic acid, arachidic acid and erucic acid all decrease, indicating lipid substances in lymph fluid are absorbed selectively after alcohol intake. It’s worth noting that arachidonic acid is closely related to inflammatory response. Furthermore, the differential metabolites are mainly related with schizophrenia, Alzheimer’s disease and lung cancer. The differential metabolites between the medium-dosage alcohol and the control group were mainly enriched in phenylalanine metabolism, valine, leucine and isoleucine biosynthesis, linoleic acid metabolism and cholesterol metabolism. The differential metabolites are mainly related to schizophrenia, Alzheimer’s disease, lung cancer and Parkinson’s disease. As the dose of alcohol increases, the contents of some metabolites in lymph fluid increase, including cholesterol, L-leucine, fumaric acid and mannitol, and the number of metabolites related to schizophrenia also tends to increase, indicating that some metabolites absorbed by the intestine-lymphatic pathway are dose-dependent on alcohol intake.Conclusion After alcohol intake, the metabolites transported via the intestinal-lymphatic pathway are significantly changed, especially in the high-dosage group. Some metabolites absorbed via the intestinal-lymphatic pathway are dose-dependent on alcohol intake. Most importantly, alcohol intake may cause inflammatory response and the occurrence of neurological diseases, psychiatric diseases and cancer diseases. High-dosage drinking may aggravate or accelerate the occurrence of related diseases. These results provide new insights into the pathogenesis of alcohol-related diseases based on the intestinal-lymphatic pathway.

Abstract:Objective Hemoglobin is the iron-containing protein in the red blood cells of many animals. The primary function of hemoglobin is to transport oxygen from lung to tissues. It is composed of two identical α-globin subunits and two identical β-globin subunits. Hemoglobin has unique magnetic properties. The paramagnetism of deoxyhemoglobin, and the diamagnetism of oxyhemoglobin and carboxyhaemoglobin have been reported previously. Studies have also shown that external magnetic field affected blood flow rate, but whether magnetic field may affect the oxygenation rate of hemoglobin remains unknown. Here in this study, we are aiming to address this question with recombinant hemoglobin. Human hemoglobin and yak hemoglobin were selected as the research objects, and a recombinant protein expression and purification system was established to explore the magnetic field effects on the oxygenation rate of hemoglobin, as well as the differences in the oxygenation rate between human hemoglobin and yak hemoglobin under external magnetic field.Methods The recombinant expression and purification system of human and yak hemoglobin was established. The recombinant hemoglobin expression was further optimized and appropriated inducing temperature and IPTG concentration were screened. Recombinant human hemoglobin and yak hemoglobin were purified to homogeneity by affinity chromatography and further by size-exclusion chromatography. SDS-PAGE was used to validate the purification, and UV-Vis spectrum and EPR were used to characterize the biochemical properties of recombinant hemoglobin. Deoxyhemoglobin of human and yak were placed under 0.3 T external magnetic field to test the magnetic field effects on oxygenation rate, and geomagnetic field condition was used as a sham control. The UV-Vis spectrum data were measured every 10 min, and the concentration and proportion of oxygenated hemoglobin, deoxyhemoglobin and methemoglobin were calculated to analyze the effects of magnetic field on the oxygenation rate of hemoglobin. The magnetic properties of human oxygenated hemoglobin and human deoxygenated hemoglobin have been measured by SQUID, a superconducting quantum interference magnetic measurement system. Three biological replications were performed for each experiment. The possible mechanism of the effect of magnetic field on the oxygenation rate of hemoglobin has been investigated and discussed.Results Human and yak hemoglobin were successfully expressed and purified by E.coli prokaryotic expression system. The optimal expression temperature was 30℃, and the most suitable IPTG concentration was 1 mmol/L. EPR results suggested that trace amount of methemoglobin existed both in the purified human hemoglobin and yak hemoglobin proteins. The oxygenation rate of yak hemoglobin appeared to be faster than that of human hemoglobin, and the additional magnetic field treatment significantly increased the oxygenation rate of both human and yak hemoglobin, and yak hemoglobin was more sensitive to magnetic field than human hemoglobin. The paramagnetism of deoxyhemoglobin was verified by SQUID measurement. However, the diamagnetism of oxygenated hemoglobin remains uncertain, probably due to the presence of trace amount of methemoglobin in the sample of oxygenated hemoglobin, which was consistent with EPR results.Conclusion In this study, human and yak hemoglobin were successfully expressed and purified. The purified hemoglobin proteins have similar function and conformational states as native protein. External static magnetic field facilitates hemoglobin oxygenation, and yak hemoglobin seems more sensitive to magnetic field compared with human hemoglobin. These findings provide theoretical basis for the potential applications of applying magnetic field to improve hypoxia symptoms in clinical practice in the future.

Abstract:Objective Spatial working memory (SWM) is an important function in cognitive behavior, and working memory impairment can seriously affect the patient’s life and cause great stress to the patient. Intermittent theta burst stimulation (iTBS) has been shown to regulate working memory function by entrainment of neural oscillations in different frequencies of the brain, but its regulation of working memory-related neural oscillations and their synchronization is not clear. The purpose of this study was to study the effect of iTBS on neural oscillation and synchronization in local and transbrain regions of rats, and to explore the mechanism of iTBS in regulating working memory.Methods Twenty-four rats were randomly divided into four groups according to their age and whether they received iTBS stimulation (AS: adult stimulation group, AC: adult control group, ES: elderly stimulation group, EC: elderly control group). Using the methods of time-frequency distribution, phase synchronization and phase-amplitude coupling analysis, the changes of local field potential signal neural oscillations in the prefrontal and hippocampal brain regions of theta and gamma bands in the process of spatial working memory behavioral tasks in each group of rats were compared and analyzed, and the relationship between the changes of neural oscillations in the two brain regions and the changes in spatial working memory ability of rats was judged based on the Pearson correlation coefficient.Results With the increase of age, the time taken by the elderly rats to learn the spatial working memory task rules increased significantly (P=0.005 6), and the time taken by iTBS stimulation to learn the SWM task rules in adult rats (P=0.001 1) and elderly rats (P=0.009 0) was shortened. At the same time, compared with adult rats, the time-frequency energy of theta and gamma band neural oscillations in the prefrontal and hippocampal brain regions of elderly rats (theta: P<0.000 1; gamma: P<0.000 1) and phase-amplitude coupling across brain regions (PFC-HPC: P=0.000 2; HPC-PFC: P=0.027 7) decreased to a certain extent, and iTBS stimulation could increase the time-frequency energy of neural oscillations of adult rats (theta: P<0.000 1; gamma: P<0.000 1) and elderly rats (theta: P=0.014 4; gamma: P=0.000 6) and the phase-amplitude coupling effect across brain regions in elderly rats (PFC-HPC: P=0.018 0; HPC-PFC: P=0.022 1). In addition, the time-frequency energy and phase-amplitude coupling of signals in each frequency band of the two brain regions were positively correlated with the behavioral accuracy of rats, while the phase synchronization of theta band and gamma band neural oscillations in the two brain regions during working memory was not correlated with the behavioral accuracy.Conclusion iTBS can enhance SWM ability and cognitive function in elderly rats, and this improvement is associated with increased coupling of time-frequency energy and cross-brain phase amplitude of neural oscillations across theta and gamma bands during SWM tasks. Similarly, in adult rats, iTBS enhances SWM ability and cognitive function by increasing the time-frequency energy of theta and gamma band neural oscillations in both brain regions during SWM tasks. Furthermore, in addition to the main findings, this study provides evidence supporting the state-dependent effects of iTBS stimulation to some extent.

Abstract:Objective Angle-closure glaucoma (ACG) is one of the major eye-blinding diseases. To diagnose ACG, it is crucial to examine the anterior chamber angle. Current diagnostic tools include slit lamp gonioscopy, water gonioscopy, ultrasound biomicroscopy (UBM), and anterior segment optical coherence tomography (AS-OCT). Slit lamp and water gonioscopy allow convenient observation of the anterior chamber angle, but pose risks of invasive operation and eye infections. UBM can accurately measure the structure of the anterior chamber angle. However, it is complex to operate and unsuitable for patients, who have undergone trauma or ocular surgery. Although AS-OCT provides detailed images, it is costly. The aim of this study is to explore a non-invasive, non-destructive optical reflection tomography (ORT) technique. This technique can achieve low-cost three-dimensional imaging and full-field anterior chamber angle measurement of the porcine eye.Methods The experiment involved assembling an optical reflection tomography system, which included a complementary metal oxide semiconductor (CMOS) camera, a telecentric system, a stepper motor, and a white light source, achieving a spatial resolution of approximately 8.5 μm. The process required positioning the porcine eye at the center of the field of the imaging system and rotating it around its central axis using a stepper motor. Reflection projection images were captured at each angle with an exposure time of 1.0 ms and an interval of 2°. The collected reflection-projection data were processed using a filtered reflection tomography algorithm, generating a series of two-dimensional slice data. These slices essentially represented cross-sectional views of the three-dimensional structural image, and were reconstructed into a complete three-dimensional structural image. Based on the reconstructed three-dimensional structural image of the porcine eye, the anterior chamber angles at different positions were measured, and a distribution map of these angles was drawn. Simultaneously, the ORT measurements were compared with the standard results obtained from optical coherence tomography (OCT) to assess the accuracy of ORT measurements.Results In this study, we successfully obtained the reflection projection data of a porcine eye using ORT technology, reconstructed its three-dimensional structural image, and measured the anterior chamber angle, generating the corresponding distribution map. To better distinguish the different structural parts of porcine eye, the three-dimensional structural image was marked with blue, green, and yellow dashed lines from the outer to the inner layers. The area between the blue and green dashed lines corresponded to the sclera. The area between the green and yellow dashed lines corresponded to the iris. The area inside the yellow dashed line corresponded to the pupil. The three-dimensional structural image clearly revealed the key anatomical features of the porcine eye. It was able to measure the anterior chamber angle at different positions. Additionally, the anterior chamber angle measurements of the porcine eye using ORT were compared with the measurements obtained using a TEL320C1 type OCT system, showing an average deviation of 0.51° and a mean square error of 0.317.Conclusion ORT is a non-invasive, non-destructive, low-cost, and high-resolution imaging technique capable of achieving three-dimensional structural imaging and full-field anterior chamber angle measurement of a porcine eye. This technology offers a new perspective for the diagnosis of angle-closure glaucoma and is significant for the screening, diagnosis, and monitoring of eye diseases, potentially benefiting clinics and small hospitals in remote areas in the future.

Abstract:Objective Detection and quantification of RNA synthesis in cells is a widely used technique for monitoring cell viability, health, and metabolic rate.After exposure to environmental stimuli, both the internal reference gene and target gene would be degraded. As a result, it is imperative to consider the accurate capture of nascent RNA and the detection of transcriptional levels of RNA following environmental stimulation. This study aims to create a Click Chemistry method that utilizes its property to capture nascent RNA from total RNA that was stimulated by the environment.Methods The new RNA was labeled with 5-ethyluridine (5-EU) instead of uracil, and the azido-biotin medium ligand was connected to the magnetic sphere using a combination of “Click Chemistry” and magnetic bead screening. Then the new RNA was captured and the transcription rate of 16S rRNA was detected by fluorescence molecular beacon (M.B.) and quantitative reverse transcription PCR (qRT-PCR).Results The bacterial nascent RNA captured by “Click Chemistry” screening can be used as a reverse transcription template to form cDNA. Combined with the fluorescent molecular beacon M.B.1, the synthesis rate of rRNA at 37℃ is 1.2 times higher than that at 15℃. The 16S rRNA gene and cspI gene can be detected by fluorescent quantitative PCR,it was found that the measured relative gene expression changes were significantly enhanced at 25℃ and 16℃ when analyzed with nascent RNA rather than total RNA, enabling accurate detection of RNA transcription rates.Conclusion Compared to other article reported experimental methods that utilize screening magnetic columns, the technical scheme employed in this study is more suitable for bacteria, and the operation steps are simple and easy to implement, making it an effective RNA capture method for researchers.