Abstract:G-protein coupled receptors (GPCRs) are an essential family of proteins on the cell membrane, widely distributed in various types of tissues and cells. Typical GPCRs are composed of characteristic 7 transmembrane α-helix domains, extracellular domain and intracellular domain. They play a key role in transmitting information inside and outside cells. These receptors can sense and respond to a variety of external signals, including odor molecules, hormones, neurotransmitters, chemokines, and so on, thereby regulating the physiological functions and metabolic activities of cells. When external signal molecules bind, these receptors undergo conformational changes, thereby activating signal transduction pathways inside cells. The most common downstream signal pathway is the activation of G proteins, but it may also activate the β-arrestin signaling pathway. This series of signal transduction processes ultimately regulates physiological processes such as cell metabolism, proliferation, and differentiation, and also plays an important role in the occurrence and development of diseases. Due to its importance in regulating cell functions and participating in the development of diseases, GPCRs have become important targets in the field of drug research and development. The mechanism of action of many drugs is achieved by intervening in the GPCR signaling pathway. As important form of function regulating, dimerization has attracted widespread attention in the research of GPCR field. In the early days, the formation of GPCR dimerization and its effect on receptor function were mainly studied by immunoprecipitation, immunofluorescence and radioligand binding experiments in overexpression systems. Nowadays, with the continuous development of biochemical and biophysical methods, more and more GPCR dimers have been identified. GPCR dimer refers to the process in which two GPCR subunits bind to each other to form a complex. The same GPCR subunits form homodimers, and different GPCR subunits form heterodimers through direct interaction. Dimerization changes the activity, affinity, internalization, localization and transport, and signal transduction characteristics of GPCR, thereby producing more complex and delicate regulation of cellular physiological processes. In recent years, the research on GPCR dimers has been continuously deepened, revealing its important role in a variety of physiological and pathological processes. In general, the structure of GPCR dimers is complex and diverse, and its formation and stability are affected by many factors, including the specificity of receptor interaction interface, the conformational changes of receptor, and the regulation of intracellular and extracellular environment. By understanding the mechanism of GPCR dimerization, we can better understand the behavior of these receptors in signal transduction and provide new ideas and opportunities for the development of novel drug targets. More and more studies have reported the dimerization of GPCR and its structure and function regulation mechanism. This article reviews the research progress on the structure and function of GPCR dimers, and summarizes some research methods and technologies, which provide a basis for understanding the discovery of GPCR dimers, dimerization methods, structure and function regulation mechanisms, and further targeting GPCR dimers. It provides a research basis for the development of polymer drugs.

Abstract:Proteins in biological systems rarely act alone, but instead bind with other biomolecules to trigger specific cellular reactions. These biomolecules are usually astonishing number of proteins self-assemble to form dimers, which are both in a relatively isolated state and in a protein interaction network and cascade. Dimerization can endow proteins with various structural and functional advantages, including improving stability, controlling the accessibility and specificity of active sites, and increasing complexity. The self-association of proteins to form dimers is a very common phenomenon, and the functional importance of homologous protein dimerization cannot be overestimated. It provides diversity and specificity in many pathways, and most cellular events, such as signal transduction, transcription cofactor recruitment, enzyme activation, and even pathogenic pathways, are significantly regulated through homologous protein-protein interactions. The regulation of protein dimerization is an important process for the growth and development of organisms under internal or external stimuli in the natural environment. Therefore, regulating the dimerization process of homologous proteins and understanding their molecular mechanisms are crucial for biomedical applications and analyzing complex biological regulatory networks. Proximity effects or physical proximity effects of molecules are essential regulatory factors in biological processes, which can be controlled through induced dimerization methods. The application range of induced proximity ranges from manipulating protein folding, activation, localization, and degradation to controlling gene transcription or cell therapy. The chemical induced dimerization (CID) system and light induced dimerization (LID) system based on proximity induction provide powerful tools for regulating the function of dimerized proteins, and have been gradually developed. The concept of CID was proposed as early as 1993. The basic principle of CID is that a small molecule controls the dimerization of a pair of proteins or domains, while binding two proteins and bringing them closer together. Small molecules in the CID system form ternary complexes with target proteins, which can bind to various sites, including “hotspot” and “allosteric sites”. Small molecules play a role by regulating protein proximity. The light induced dimerization system uses photosensitive proteins to undergo conformational changes under light, thereby inducing protein interactions. Multiple photosensitive proteins derived from plants and microorganisms can undergo photo induced homologous interactions, and relying on LID systems, they can be used to study various biological processes, including cell signal transduction, microbial synthesis, and biomedical applications. In recent years, metal ions, nucleic acids, and molecular host guest systems have been proposed as new methods for orthogonal control of homologous protein dimerization, expanding the development and application of dimerization systems. In addition, the chemo-optogenetic approach combines the advantages of CID and LID systems and has also been applied in inducing protein dimerization. This review elaborates on the methods and applications of inducing homodimerization of proteins through CID system, LID system, and supramolecular chemistry, while discussing the advantages and disadvantages of dimerization systems. The development direction of dimerization systems is also discussed, in order to provide some reference and ideas for the future application and development of homologous protein dimerization.

Abstract:The dynamin superfamily protein (DSP) encompasses a group of large GTPases that are involved in various membrane remodeling processes within the cell. These proteins are characterized by their ability to hydrolyze GTP, which provides the energy necessary for their function in membrane fission, fusion, and tubulation activities. Dynamin superfamily proteins play critical roles in cellular processes such as endocytosis, organelle division, and vesicle trafficking. It is typically classified into classical dynamins and dynamin-related proteins (Drp), which have distinct roles and structural features. Understanding these proteins is crucial for comprehending their functions in cellular processes, particularly in membrane dynamics and organelle maintenance. Classical dynamins are primarily involved in clathrin-mediated endocytosis (CME), a process crucial for the internalization of receptors and other membrane components from the cell surface into the cell. These proteins are best known for their role in pinching off vesicles from the plasma membrane. Structually, classical dynamins are composed of a GTPase domain, a middle domain, a pleckstrin homology (PH) domain that binds phosphoinositides, a GTPase effector domain (GED), and a proline-rich domain (PRD) that interacts with SH3 domain-containing proteins. Functionally, the classical dynamins wrap around the neck of budding vesicles, using GTP hydrolysis to constrict and eventually acting as a “membrane scissor” to cut the vesicle from the membrane. In mammals, there are three major isoforms: dynamin 1 (predominantly expressed in neurons), dynamin 2 (ubiquitously expressed), and dynamin 3 (expressed in testes, lungs, and neurons). Recent studies have also revealed some non-classical functions of classical dynamins, such as regulating the initiation and stabilization of clathrin-coated pits (CCPs) at the early stages of CME, influencing the formation of the actin cytoskeleton and cell division. Drps share structural similarities with classical dynamins but are involved in a variety of different cellular processes, primarily related to the maintenance and remodeling of organelles, and can be mainly categorized into “mediating membrane fission”, “mediating membrane fusion” and “non-membrane-dependent functions”. Proteins like Drp1 are crucial for mitochondrial division, while others like Fis1, Mfn1, and Mfn2 are involved in mitochondrial and peroxisomal fission and fusion processes, which are essential for the maintenance of mitochondrial and peroxisomal integrity and affect energy production and apoptosis. Proteins like the Mx protein family exhibit antiviral properties by interfering with viral replication or assembly, which is critical for the innate immune response to viral infections. Some other proteins are involved in the formation of tubular structures from membranes, which is crucial for the maintenance of organelle morphology, particularly in the endoplasmic reticulum and Golgi apparatus. Studies on dynamin superfamily proteins have been extensive and have significantly advanced our understanding of cellular biology, disease mechanisms, and therapeutic potential. These studies encompass a broad range of disciplines, including molecular biology, biochemistry, cell biology, genetics, and pharmacology. By comprehensively summarizing and organizing the structural features and functions of various members of the dynamin superfamily protein, this review not only deepens the understanding of its molecular mechanisms, but also provides valuable insights for clinical drug research related to human diseases, potentially driving further advancements in the field.

Abstract:Tight-junction (TJ) is a complex supramolecular entity composed of complete membrane proteins, membranes and soluble cytoplasmic proteins, which is distributed in almost all barrier structures in the body. It can maintain the polarity of epithelial cells, close the intercellular space and prevent the overflow of materials in the epithelial space, and is a highly dynamic signaling entity. Occludin is one of the most representative members of TJ proteins, mainly responsible for sealing intercellular connections, maintaining intercellular permeability, and participating in maintaining the integrity of vascular endothelium. The integrity of occludin is related to the integrity of TJ, and the function of occludin is often associated with the barrier properties of various tissues, and the abnormal expression of occludin is related to the occurrence and development of various diseases. Occludin contains abundant Ser and Thr residues and has multiple phosphorylation sites. Phosphorylation is necessary for the combination of occludin and TJ, which can regulate the location of occludin, regulate the expression of occludin, and enhance the permeability and barrier function of TJ. Therefore, phosphorylation regulation is a mechanism that cannot be ignored in the regulation of occludin function. Occludin also interacts with many other proteins, such as co-forming the cytoskeleton with ZO-1, and is regulated by a variety of transcription factors. Studies have confirmed that in pathological conditions, a variety of signaling pathways can disrupt the integrity of cell barrier by regulating the expression and distribution of occludin. Myosin light chain kinase (MLCK) signal transduction pathway is one of the important ways to regulate the structure and function of TJ. It influences the expression of occludin by altering the cytoskeleton. MLCK mainly uses the phosphorylation of myosin light chain (MLC) as a medium to promote actin contraction, secondary decomposition of tightly binding proteins, resulting in increased or changed cellular barrier permeability, and increased MLC phosphorylation is also a biochemical marker of actomyosin contraction. Activation of MLCK causes Thr18 and Ser19 phosphorylation of MLC, which promotes the assembly of myosin II into myosin fibers and activates the hydrolysis of ATP, which relaxes the intercellular connections and reduces the ability of upper cortex to resist external invaders. Protein kinase C (PKC) plays an important role in the regulation of tightly connected signaling molecules, affecting the dynamic changes of paracellular permeability. PKC pathway is a key link in many cell signal transduction pathways, which influences all aspects of cell activities by catalyzing Ser/Thr residues phosphorylation of membrane proteins and many enzyme proteins. After PKC activation, it can regulate cellular barrier function by phosphorylating occludin and inducing its redistribution, and directly affect TJ action. Specific PKC subunits such as PKCα, PKCδ and PKCγ are activated and act on occludin molecules to promote their phosphorylation and cause the increase of TEER. The increase of TEER helps to regulate intercellular TJ and enhance the tightness of intercellular connections. Mitogen-activated protein kinases (MAPK) are usually activated by inflammatory factors, during which different signal transduction pathway subfamilies are formed to regulate occludin expression and affect tight junction and mucosal barrier functional integrity. Meanwhile, occludin is easily affected by various factors (such as cytokines and flora toxins), and abnormal expression of occludin will lead to structural damage of TJ and further damage of the intercellular barrier. Therefore, this paper summarizes the molecular structure and physiological function of occludin, and further summarizes its related signal regulation pathways and influencing factors, in order to provide theoretical support for maintaining the integrity of barrier function of occludin.

Abstract:Isocitrate dehydrogenase 1 (IDH1) R132H is the most common mutated gene in grade II-III gliomas and oligodendrogliomas. Instead of activating telomerase (a reverse transcriptase which using RNA as a template to extend telomere length), the majority of IDH1^R132H mutant glioma maintain telomere length through an alternative mechanism that relies on homologous recombination (HR), which is known as alterative lengthening of telomere (ALT).The phenotype of ALT mechanism include: ALT associated promyelocytic leukemia protein (PML) bodies (APBs); extrachromosomal telomeric DNA repeats such as C- and T-loops; telomeric sister chromatid exchange (T-SCE), etc. The mechanism of ALT activation is not fully understood. Recent studies have shown that mutation IDH1 contributes to ALT phenotype in glioma cells in at least three key ways. Firstly, the IDH1^R132H mutation mediates RAP1 down-regulation leading to telomere dysfunction, thus ensuring persistent endogenous telomeric DNA damage, which is important for ALT activation. Spontaneous DNA damage at telomeres may provide a substrate for mutation break-induced replication (BIR)-mediated ALT telomere lengthening, and it has been demonstrated that RAP1 inhibits telomeric repeat-containing RNA, transcribed from telomeric DNA repeat sequences (TERRA) transcription to down-regulate ALT telomere DNA replication stress and telomeric DNA damage, thereby inhibiting ALT telomere synthesis. Similarly, in ALT cells, knockdown of telomere-specific RNaseH1 nuclease triggers TERRA accumulation, which leads to increased replication pressure. Overexpression of RNaseH1, on the other hand, attenuates the recombination capacity of ALT telomeres, leading to telomere depletion, suggesting that RAP1 can regulate the level of replication pressure and thus ALT activity by controlling TERRA expression. Secondly, the IDH1^R132H also alters the preference of the telomere damage repair pathway by down-regulating XRCC1, which inhibits the alternative non-homologous end joining (A-NHEJ) pathway at telomeres and alters cellular preference for the HR pathway to promote ALT. Finally, the IDH1^R132H has a decreased affinity for isocitric acid and NADP+ and an increased affinity for α ketoglutarate (α-KG) and NADPH, so that the mutant IDH1^R132H catalyzes the hydrogenation of α-KG to produce 2-hydroxyglutarate (2-HG)in a NADPH-dependent manner. Because 2-HG is structurally similar to α-KG, which maintains the trimethylation level of H3k9me3 by competitively inhibiting the activity of the α-KG-dependent histone demethylase KDM4B, and recruits heterochromatin protein HP1α to heterochromatinize telomeres, and promote ALT phenotypes in cooperation with the inactivating of ATRX. In addition, it has been shown that APBs contain telomeric chromatin, which is essentially heterochromatin, and HP1α is directly involved in the formation of APBs. Based on these studies, this article reviews the mechanism of IDH1^R132H mediated telomere dysfunction and the preference of DNA repair pathway at telomeres in cooperate with ATRX loss to promote ALT, which may provide references for clinical targeted therapy of IDH1^R132H mutant glioma.

Abstract:In recent years, the development of host-acting antibacterial compounds has gradually become a hotspot in the field of anti-infection. Through research on the interaction mechanism between hosts and pathogenic bacteria, it has been found that the immune system is one of the key targets of host-acting antibacterial compounds. There is a communication system called the quorum sensing system in microorganisms, which mainly adjusts the structure of multi-microbial community and coordinates the group behavior. When the quorum sensing molecules secreted by microorganisms reach a threshold concentration, the quorum sensing system is activated and the overall gene expression of the microorganism is changed. In addition to regulating the density of microorganisms, quorum sensing molecules can also act as a link between pathogenic microorganisms and hosts, entering the host immune system and playing a role in affecting the morphological structure of immune cells, secreting cytokines, and inducing apoptosis, leading to host immune injury and causing host immune dysfunction.The key mechanism of 3-oxo-C12-HSL and other acyl-homoserine lactone (AHL) molecules in the innate immune system has been extensively studied. The lipid solubility allows AHLs to pass through the plasma membrane of host immune cells easily and induce dissolution of lipid domains. Then, it acts through signaling pathways such as p38MAPK and JAK-STAT, further influencing the immune cell’s defense response to bacteria and potentially leading to cell apoptosis. Additionally, the human lactonase paraoxonase 2, which can degrade 3-oxo-C12-HSL, has been found in macrophage. It acts as an immune regulator that promotes macrophage phagocytosis of pathogens and is hypothesized to have the ability to reduce bacterial resistance. The mechanism of quorum sensing molecules in the adaptive immune system is less studied, the current results suggest that 3-oxo-C12-HSL is closely related to the mitochondrial pathway in host immune cells. For example, 3-oxo-C12-HSL induces apoptosis of Jurkat cells by inhibiting the expression of three mitochondrial electron transport chain proteins; it can also trigger mitochondrial dysfunction and induce mast cell apoptosis through Ca²⁺ signaling.Among the quorum sensing molecules, the AHLs have the greatest impact on plant immune system. The different effects on plant resistance depends on the chain lengths of acyl groups in bacterial-produced AHLs. Short-chain AHLs (C4-HSL and C8-HSL) induce plant resistance to pathogenic bacteria mainly through the auxin pathway and jasmonic acid pathway. Long-chain AHL (3-oxo-C14-HSL) is commonly used in hosts against fungal pathogens by inducing stomata defense responses, and the reaction process is related to salicylic acid. Diffusible signal factor molecules also interfere with the stomatal immunity caused by pathogens. It may act through the formin nanoclustering-mediated actin assembly and MPK3 pathway to inhibit the innate immunity of Arabidopsis. In summary, AHLs induced different plant pathways and affects the plant-bacteria interactions to trigger plant immunity. As a quorum sensing molecule of fungi, farnesol has similar effects on host immunity as AHLs, such as stimulating cytokine secretion and activating an inflammatory response. It also plays a unique role on dendritic cell differentiation and maturation. In addition, studies have found that farnesol has a protective effect on autoimmune encephalomyelitis, which may be related to its effect on the composition of intestinal microorganisms of the host.Therefore, targeting the host immune system and quorum sensing molecules to develop antibacterial compounds can effectively inhibit the invasion of pathogens and subserve the host to resist the influence of pathogenic bacteria. This article will review the mechanism of host immune responses triggered by important quorum sensing molecules, aiming to explore the targets of host-acting antibacterial compounds and provide new directions for the prevention or treatment of causative infectious sources and the development of related drugs.

Abstract:Cyclin-dependent kinases (CDKs) are proline-induced serine/threonine kinases that are primarily involved in the regulation of cell cycle, gene transcription, and cell differentiation. In general, CDKs are activated by binding to specific regulatory subunits of cell cycle proteins and are regulated by phosphorylation of specific T-loops by CDK activated kinases. In the CDKs family, cyclin-dependent kinase 5 (CDK5) is a specialized member whose activity is triggered only by interaction with p35 and p39, which do not have the same sequence as the cell cycle proteins, and this may be one reason why CDK5 is distinguished from other CDK members by its structural and functional differences. In addition, unlike most CDK members that require phosphorylation at specific sites to function, CDK5 does not require such phosphorylation, and it can be activated simply by binding to p35 and p39. More notably, inhibitors that are commonly used to inhibit the activity of other CDK members have almost zero effect on CDK5. In contrast, CDK5, as a unique CDK family member, plays an important role in the development of numerous diseases. In metabolic diseases, elevated CDK5 expression leads to decreased insulin secretion, increased foam cell formation and triggers decreased bone mass in the body, thus accelerating metabolic diseases, and the role of CDK5 in bone biology is gradually gaining attention, and the role of CDK5 in bone metabolic diseases may become a hotspot for research in the future; in neurodegenerative diseases, hyperphosphorylation of Tau protein is an important hallmark of Alzheimer’s disease development, and changes in CDK5 expression are associated with Tau protein phosphorylation and nerve death, indicating that CDK5 is highly related to the development of the nervous system; in tumor diseases, the role of CDK5 in the proliferation, differentiation and migration and invasion of tumor cells marks the development of tumorigenesis, but different researchers hold different views, and further studies are needed in the follow-up. Therefore, the study of its mechanism of action in diseases can help to reveal the pathogenesis and pathological process of diseases. Appropriate exercise not only helps in the prevention of diseases, but also plays a positive role in the treatment of diseases. Exercise-induced mechanical stress can improve bone microstructure and increase bone mass in osteoporosis patients. In addition, exercise can effectively inhibit neuronal apoptosis and improve mitochondrial dysfunction, more importantly, appropriate exercise can inhibit the proliferation of cancer cells to a certain extent. It can be seen that exercise occupies a pivotal position in the prevention and treatment of pathologic diseases. It has been shown that exercise can reduce the expression of CDK5 and affect the pathological process of neurological diseases. Currently, there is a dearth of research on the specific mechanisms of CDK5’s role in improving disease outcomes through exercise. In order to understand its effects more comprehensively, subsequent studies need to employ diverse exercise modalities, targeting patients with various types of diseases or corresponding animal models for in-depth exploration. This article focuses on the pathological functions of CDK5 and its relationship with exercise, with a view to providing new insights into the prevention and treatment of disease by CDK5.

Abstract:Parkinson’s disease (PD) is a neurodegenerative disorder characterized by muscle rigidity, resting tremor, and postural instability, which severely impair the quality of life in middle-aged and elderly individuals. PD’s pathogenesis is complex, involving oxidative stress, immune inflammation, and genetic factors. Despite extensive research, precise therapeutic targets for PD remain elusive, necessitating further investigation into its underlying mechanisms. Recent studies highlight the pivotal role of regional brain iron overload, oxidative stress, and lipid peroxidation in PD’s pathogenesis. Ferroptosis, a form of regulated cell death driven by iron dependency and lipid peroxidation, has emerged as a critical factor in PD pathology. This review examines the relationship between ferroptosis and PD and explores the potential of exercise as a therapeutic intervention to modulate ferroptosis and alleviate PD symptoms. Ferroptosis, distinct from other forms of cell death such as necrosis, autophagy, pyroptosis, and apoptosis, is characterized by mitochondrial shrinkage, reduced cristae, and membrane collapse, without nuclear fragmentation, DNA cleavage, or caspase activation. It is induced by the accumulation of intracellular Fe²⁺, which enhances lipid peroxidation and reactive oxygen species (ROS) generation, ultimately leading to cell death. Studies show disrupted iron metabolism in PD patients, with elevated iron levels in dopaminergic neurons of the substantia nigra correlating with disease severity. Iron chelation therapy has shown promise in alleviating PD symptoms by reducing brain iron levels, highlighting the significance of iron metabolism in PD pathogenesis. Lipid peroxidation, a hallmark of ferroptosis, involves the oxidation of polyunsaturated fatty acids (PUFAs) in cell membranes, compromising membrane integrity and increasing permeability. Elevated lipid peroxidation in the substantia nigra contributes to neuronal damage in PD. Enzymes such as ACSL4 and LPCAT3, crucial in PUFA metabolism, play significant roles in ferroptosis. Exercise has been shown to modulate these enzymes, potentially reducing lipid peroxidation and preventing ferroptosis in PD. Glutathione (GSH) metabolism is another crucial factor in ferroptosis regulation. GSH depletion impairs ROS detoxification, exacerbating oxidative stress and lipid peroxidation. PD patients exhibit reduced GSH levels in the substantia nigra, making dopaminergic neurons more vulnerable to oxidative damage. Exercise enhances GSH synthesis and activity, mitigating oxidative stress and ferroptosis in PD. α-Synuclein aggregation, a hallmark of PD, is closely linked to iron metabolism and oxidative stress. Excessive α-synuclein binds to iron, promoting its aggregation and inducing ferroptosis. Exercise has been found to reduce α-synuclein accumulation and its pathological phosphorylation, potentially through the upregulation of neuroprotective proteins like DJ-1 and Irisin. These proteins enhance antioxidant defenses and facilitate α-synuclein degradation, providing a protective effect against PD progression. Additionally, glutamate excitotoxicity, driven by dysregulated glutamate metabolism and receptor activity, contributes to ferroptosis in PD. Exercise modulates glutamate levels and receptor expression, reducing excitotoxicity and iron-induced neuronal damage. In conclusion, emerging research suggests that exercise may inhibit ferroptosis through multiple mechanisms, including regulation of iron metabolism, enhancement of antioxidant defenses, reduction of α-synuclein aggregation, and modulation of glutamate metabolism. These findings highlight the potential of exercise as a non-pharmacological intervention in the prevention and treatment of PD. Further research is needed to elucidate precise mechanisms and optimize exercise protocols for maximum therapeutic benefit.

Abstract:As the global population continues to age, the incidence of Alzheimer’s disease (AD), one of the most common neurodegenerative diseases, continues to rise significantly. As the disease progresses, the patient’s daily living abilities gradually decline, potentially leading to a complete loss of self-care abilities. According to estimates by the Alzheimer’s Association and the World Health Organization, AD accounts for 60%-70% of all other dementia cases, affecting over 55 million people worldwide. The case number is estimated to double by 2050. Despite extensive research, the precise etiology and pathogenesis of AD remain elusive. Researchers have a profound understanding of the disease’s pathological hallmarks, which include amyloid plaques and neurofibrillary tangles resulting from the abnormal phosphorylation of Tau protein. However, the exact causes and mechanisms of the disease are still not fully understood, leaving a vital gap in our knowledge and understanding of this debilitating disease. A crucial player that has recently emerged in the field of AD research is the α7 nicotinic acetylcholine receptor (α7nAChR). α7nAChR is composed of five identical α7 subunits that form a homopentamer. This receptor is a significant subtype of acetylcholine receptor in the central nervous system and is widely distributed in various regions of the brain. It is particularly prevalent in the hippocampus and cortical areas, which are regions associated with learning and memory. α7nAChR plays a pivotal role in several neurological processes, including neurotransmitter release, neuronal plasticity, cell signal transduction, and inflammatory response, suggesting its potential involvement in numerous neurodegenerative diseases, including AD. In recent years, the role of α7nAChR in AD has been the focus of extensive research. Emerging evidence suggests that α7nAChR is involved in several critical steps in the disease progression of AD. These include involvement in the metabolism of amyloid β-protein (Aβ), the phosphorylation of Tau protein, neuroinflammatory response, and oxidative stress. Each of these processes contributes to the development and progression of AD, and the involvement of α7nAChR in these processes suggests that it may play a crucial role in the disease’s pathogenesis. The potential significance of α7nAChR in AD is further reinforced by the observation that alterations in its function or expression can have significant effects on cognitive abilities. These findings suggest that α7nAChR could be a promising target for therapeutic intervention in AD. At present, the results of drug clinical studies targeting α7nAChR show that these compounds have improvement and therapeutic effects in AD patients, but they have not reached the degree of being widely used in clinical practice, and their drug development still faces many challenges. Therefore, more research is needed to fully understand its role and to develop effective treatments based on this understanding. This review aims to summarize the current understanding of the association between α7nAChR and AD pathogenesis. We provide an overview of the latest research developments and insights, and highlight potential avenues for future research. As we deepen our understanding of the role of α7nAChR in AD, it is hoped that this will pave the way for the development of novel therapeutic strategies for this devastating disease. By targeting α7nAChR, we may be able to develop more effective treatments for AD, ultimately improving the quality of life for patients and their families.

Abstract:Traumatic spinal cord injury (SCI) refers to damage to the structure and function of spinal cord caused by external trauma. This damage results in the loss of sensation, movement, or autonomous functions, which can lead to partial or complete paralysis and impact the patients’ independence and quality of life. Studying drugs related to spinal cord injuries and their mechanisms of action will help enhance patients’ quality of life and alleviate social and economic burdens. Traumatic spinal cord injury can be categorized into primary and secondary injuries. It leads to ongoing neurodegeneration, inflammation, and scarring, necessitating continuous intervention to reduce the cascading effects of secondary injuries. Regenerative repair of SCI has been one of the most challenging problems in medicine. It is characterized by the involvement of microglia, phagocytes (including neutrophils and monocytes), and antigen-presenting cells of the central nervous system, such as dendritic cells. These inflammatory mediators contribute to axonal demyelination and degeneration, leading to severe nerve damage. Currently, there has been little progress in the clinical treatment of SCI. Current clinical modalities, such as surgical interventions and hormone shock therapies, have not yielded specific pharmacotherapeutic options, hindering significant functional recovery. The current treatment methods are ineffective in alleviating oxidative stress and neuroinflammatory responses caused by spinal cord injury. They also do not offer neural protection, resulting in ongoing neurofunctional degradation. Intravenous injection of methylprednisolone through the arm has been used as a treatment option for spinal cord injury. Recent studies have shown that the potential side effects of the drug, such as blood clots and pneumonia, outweigh its benefits. Methylprednisolone is no longer recommended for the routine treatment of spinal cord injury. In recent years, significant progress has been made in spinal cord injury intervention through the use of nanotechnology and biomaterials. Nanozymes can enhance the therapeutic efficacy of spinal cord injury by catalyzing the clearance of free radicals similar to enzymes and suppressing inflammatory responses. Nanozymes can reduce the degree of fibrosis, promote neuron survival and angiogenesis, and provide favorable conditions for tissue regeneration. Through in vitro and in vivo toxicology experiments, it was found that the nanozyme demonstrates good biocompatibility and safety. It did not cause any significant changes in body weight, hematological indicators, or histopathology. These findings indicate the potential for its clinical applications. Based on current research results and discoveries, nanozymes have broad application prospects in the biomedical field. There are numerous potential research directions and application areas that are worthy of further exploration and development. Although there have been preliminary studies on the catalytic performance of nanozymes, further research is needed to thoroughly investigate their catalytic mechanisms. Further exploration of the interaction between nanozymes and substrates, reaction kinetics, and factors affecting catalytic activity will help to better understand their mechanism of action in the field of biocatalysis.

Abstract:As a microbial therapy method, fecal microbiota transplantation (FMT) has attracted the attention of researchers in recent years. As one of the most direct and effective methods to improve gut microbiota, FMT achieves therapeutic benefits by transplanting functional gut microbiota from healthy human feces into the intestines of patients to reconstruct new gut microbiota. FMT has been proven to be an effective treatment for gastrointestinal diseases such as Clostridium difficile infection, irritable bowel syndrome, and inflammatory bowel disease. In addition, the clinical and basic research of FMT outside the gastrointestinal system is also emerging. It is worth noting that there is bidirectional communication between the gut microbial community and the central nervous system (CNS) through the gut-brain axis. Some gut bacteria can synthesize and release neurotransmitters such as glutamate, gamma-aminobutyric acid (GABA) and dopamine. Imbalanced gut microbiota may interfere with the normal levels of these neurotransmitters, thereby affecting brain function. Gut microbiota can also produce metabolites that may cross the blood-brain barrier and affect CNS function. FMT may affect the occurrence and development of CNS and its related diseases by reshaping the gut microbiota of patients through a variety of pathways such as nerves, immunity, and metabolites. This article introduces the development of FMT and the research status of FMT in China, and reviews the basic and clinical research of FMT in neurodegenerative diseases (Alzheimer’s disease, Parkinson’s disease), neurotraumatic diseases (spinal cord injury, traumatic brain injury) and stroke from the characteristics of three types of nervous system diseases, the characteristics of intestinal flora, and the therapeutic effect and mechanism of fecal microbiota transplantation, summarize the common mechanism of fecal microbiota transplantation in the treatment of CNS diseases and the therapeutic targets. We found that the common mechanisms of FMT in the treatment of nervous system diseases may include the following 3 categories through summary and analysis. (1) Gut microbiota metabolites, such as SCFAs, TMAO and LPS. (2) Inflammatory factors and immune inflammatory pathways such as TLR-MyD88 and NF-κB. (3) Neurotransmitter 5-HT. In the process of reviewing the studies, we found the following problems. (1) In basic researches on the relationship between FMT and CNS diseases, there are relatively few studies involving the autonomic nervous system pathway. (2) Clinical trial studies have shown that FMT improves the severity of patients’ symptoms and may be a promising treatment for a variety of neurological diseases. (3) The improvement of clinical efficacy is closely related to the choice of donor, especially emphasizing that FMT from healthy and young donors may be the key to the improvement of neurological diseases. However, there are common challenges in current research on FMT, such as the scientific and rigorous design of FMT clinical trials, including whether antibiotics are used before transplantation or different antibiotics are used, as well as different FMT processes, different donors, different functional analysis methods of gut microbiota, and the duration of FMT effect. Besides, the safety of FMT should be better elucidated, especially weighing the relationship between the therapeutic benefits and potential risks of FMT carefully. It is worth mentioning that the clinical development of FMT even exceeds its basic research. Science and TIME rated FMT as one of the top 10 breakthroughs in the field of biomedicine in 2013. FMT therapy has great potential in the treatment of nervous system diseases, is expected to open up a new situation in the medical field, and may become an innovative weapon in the medical field.

Abstract:Osteoscarcopenia (OS) is a common degenerative syndrome in the elderly, which is caused by a decrease in both bone and muscle mass during the aging process, leading to osteoporosis and sarcopenia, a decrease in body balance, and a risk of falls and fractures, posing a serious threat to the quality of life and lifespan of the elderly. Osteoskeletal dystrophy increases with age, and its occurrence is higher in females than that in males. At present, there is no unified diagnostic standard, making it impossible to achieve early detection and intervention. The commonly used diagnostic methods include quantitative computed tomography (CT), magnetic resonance imaging (MRI), dual energy X-ray absorptiometry (DXA), muscle mass bioelectrical impedance analysis (BIA), as well as daily gait speed (UGS), short physical performance battery (SPPB), timed start test (TUG), and biochemical evaluation indicators to improve early diagnosis and screening. Due to the fact that both bones and muscles belong to the motor system, osteoporosis and sarcopenia share common pathogenic factors in genetics, endocrine, paracrine, and fat infiltration, which interact and regulate each other, inducing the occurrence of osteoscarcopenia. Osteoporosis and sarcopenia, two age-related diseases, share the same pathogenesis and regulatory pathways, as well as common drug targets. For example: somatostatin α-actin-3, peroxisome proliferator activated receptor γ coactivation factor-1α (PGC-1α), myocyte enhancer factor-2 (MEF2C), sterol regulatory element binding transcription factor 1 (SREBF1), protoadhesion 7 (PCDH7) and methyltransferase like 21C (METTL21C), osteocalcin and bone derived bone factor gap junction connexin 43 (Cx43), growth hormone (GH), sex hormones, and diseases (such as tumors, diabetes, polycystic ovary syndrome, cardiovascular disease, anemia, disability, inflammatory disease), aging, nutrition, and poor living habits are closely related to osteosarcopenia. Osteoporosis is characterized by low bone mass and microstructural degeneration of bone tissue, while sarcopenia is characterized by loss of muscle mass, strength, and function, both of which often coexist in the elderly population. Exercise regulates muscle and skeletal cytokines such as myostatin (MSTN) and irisin β-aminoisobutyric acid (BAIBA), brain derived neutrophil factor (BDNF), interleukin, prostaglandin E2, Wnt, osteocalcin (OCN), and transforming growth factor-β (TGF-β) and receptor activator of NF-κB ligand (RANKL) interfere with each other to prevent and treat osteoscarcopenia. Wnt/β-catenin signaling pathway can simultaneously regulate the growth and metabolism of bones and muscles, and promote osteoblast proliferation, maturation, and mineralization by increasing OPG/RANKL, which is beneficial for bone mass increase and induces proliferation of muscle satellite cells, stimulating and promoting increased muscle synthesis. NF-κB pathway is the main regulatory factor for inflammation mediated muscle atrophy. Meanwhile, NF-κB DNA can participate in RANKL inducing osteoclast differentiation in bone tissue, thereby reducing bone mass. Although exercise and nutrition can improve the symptoms of osteoporosis, they cannot be completely cured, and there are no specific drugs in clinical practice that can cure sarcopenia. Because osteoscarcopenia has a common crosstalk mechanism in the aging process, it is of great significance to prevent osteoscarcopenia by improving bone mass and muscle content through exercise, nutrition, and medication.

Abstract:Plant natural products have a wide range of pharmacological properties, not only can they be used as plant dietary supplements to meet the nutritional needs of the human body in the accelerated pace of life, but also occupy an important position in the research and development of therapeutic drugs for the treatment of tumors, inflammation and other diseases, and have been widely accepted by the public due to their good safety. However, despite the above advantages of plant natural products, limiting factors such as low solubility, poor stability, lack of targeting, high toxicity and side effects, and unacceptable odor have greatly impeded their conversion to clinical applications. Therefore, the development of new avenues for the application of new natural products has become an urgent problem to be solved at present. In recent years, with the continuous development of research, various strategies have been developed to improve the bioavailability of natural products. Among them, nanocarrier delivery system is one of the most attractive strategies at present. In past studies, a large number of nanomaterials (organic, inorganic, etc.) have been developed to encapsulate plant-derived natural products for their efficient delivery to specific organs and cells. Up to now, nanotechnology has not only been limited to pharmaceutical applications, but is also competing in the fields of nanofood processing technology and nanoemulsions. Among the various nanocarriers, liposomes are the largest nanocarriers with the largest market share at present. Liposomes are bilayer nanovesicles synthesized from amphiphilic substances, which have advantages such as high drug loading capacity and stability. Attractively, the flexible surface of liposomes can be modified with various functional elements. Functionalized modification of liposomes with different functional elements such as antibodies, nucleic acids, peptides, and stimuli-responsive moieties can bring out the excellent drug delivery function of liposomes to a greater extent. For example, the modification of functional elements with targeting function such as nucleic acids and antibodies on the surface of liposomes can deliver natural products to the target location and improve the bioavailability of drugs; the modification of stimulus-responsive groups such as photosensitizers, magnetic nanoparticles, pH-responsive groups, and temperature sensitizers on the surface of liposomes can achieve controlled release of drugs, localized targeting, and synergistic thermotherapy. In addition to the above properties, by using functionalized liposomes to encapsulate natural products with irritating properties can also effectively mask the irritating properties of natural products, improve public acceptance, and increase the possibility of application of irritating natural products. There are various strategies for modifying liposomes with functional elements, and the properties of functionalized liposomes constructed by different construction strategies differ. The commonly used construction strategies for functionalized liposomes include covalent modification and non-covalent modification. These two types of construction strategies have their own advantages and disadvantages. Covalent modification has better stability than non-covalent modification, but its operation is cumbersome. With the above background, this review focuses on the three typical problems faced by plant natural products at present, and summarizes the specific applications of functionalized liposomes in them. In addition, this paper summarizes the construction strategies for building different types of functionalized liposomes. Finally, this paper will also review the opportunities and challenges faced by functionalized liposomes to enter clinical therapy, and explore the opportunities to overcome these problems, with a view to better realizing the precise control of plant nanomedicines, and providing ideas and inspirations for researchers in related fields as well as relevant industrial staff.

Abstract:Radioactive contamination can occur during nuclear accidents, loss of radioactive sources and the use of radiation for photography, disinfection and detection. When the human body is accidentally contaminated by radionuclides, radionuclides can cause harm to the human body through inhalation, ingestion, direct transdermal absorption and contaminated wounds into body tissues and organs. In the treatment of radionuclide contamination in vivo, the main way is decorporation therapy, which mainly uses specific decorporation agents to selectively bind radionuclides to form stable non-toxic complexes, thereby preventing their deposition in the body, accelerating excretion, and reducing the total accumulation of radionuclides in human tissues. At present, internal radionuclide decorporation agents promote the release of radionuclides from the body mainly by stopping the entry of radionuclides into the body, ion exchange, chelation, and binding of exportants to carriers. But recent studies have found that lysosomal exocytosis, the natural clearing function of activated cells, also has a significant exportation effect. In this paper, we first introduced and analyzed the mechanism and research status of radionuclide decorporation agents that have been used in clinical practice, such as the blocking effect of potassium iodide, the ion exchange effect of Prussian blue, the chelation effect of DTPA, and the urine alkalinization effect of sodium bicarbonate. The second part introduces the mechanism and research status of promising radionuclide decorporation agents. Among them, 3,4,3-LI (1,2-HOPO) and 5-LIO (Me-3,2-HOPO) are the most promising ones and have been approved for phase I clinical trials. Others such as catecholamines, polyethyleneimine and fullerenes are also being studied with great potential. Polyethyleneimine, as a biological macromolecular chelator, has more chelating sites and stronger targeting effects than small molecule chelators, and has achieved a real breakthrough in decorporation. Fullerenes are known as “free radical sponges” with good free radical scavenging ability and antioxidant properties. In recent years, biomaterials have been widely used in the field of radionuclide decorporation, which has greatly improved the decorporation efficiency. Chitosan and pectin have shown great advantages in promoting radionuclide decorporation, chitosan can adsorb metal ions through electrostatic interaction and chelation, and can also react with free radicals to remove free radicals generated after radionuclides enter the body. Pectin can promote uranium efflux, but the exact mechanism remains unclear. Liposomes and nanomaterials as carriers enhance the intracellular drug delivery, prolong the retention time of drugs in the body, reduce adverse reactions, and make the traditional efflux enhancers glow with new vitality and have good development prospects. The last part summarizes and looks forward to the future research direction of radionuclide decorporation agents. At present, the research on decorporation agents at home and abroad is mostly stuck in the stage of drug development and drug synthesis, and few have actually entered the clinical trial stage. Therefore, the optimization of existing decorporation agents and the development of new ligands are critical. The targeting, biological safety, oral availability, and treatment needs of large-scale contamination scenarios are still the focus of attention. In addition, from the point of view of the mechanism itself, it is a new idea to promote the emission of radionuclides by activating potential channels, which can be continuously explored.

Abstract:Objective GQDs has become a superstar among zero-dimensional carbon-based materials. As one of the most abundant and important biological elements, its unique optical properties, high dispersion and biocompatibility have attracted extensive attention from scientists. This paper aims to investigate the effect of GQDs on cell viability, apoptosis and inflammatory factor expression in RAW264.7 macrophages and evaluate cell imaging capability of GQDs in vitro, which could provide theoretical basis for the safe application of GQDs in biomedical field.Methods Graphene oxide was prepared by modified Hummer’s method. H₂O₂ and W₁₈O₄₉ interacted with each other under hydrothermal conditions to produce hydroxyl radicals, which can cut graphene oxide into GQDs using a top-down approach. The microstructure of GQDs was analyzed in detail by X-ray powder diffraction, X-ray photoelectron spectroscopy, transmission electron microscopy, atomic force microscopy, scanning electron microscopy and Fourier infrared transform. The biocompatibility of GQDs on macrophage was evaluated by CCK-8 and dead/alive staining. Flow cytometry results showed the apoptosis of RAW264.7 macrophages induced by GQDs. mRNA expression of inflammatory factors was evaluated by RT-qPCR. Cell imaging is exhibited by laser scanning confocal.Results Hydroxyl radicals are produced by H₂O₂ and W₁₈O₄₉ under hydrothermal conditions, which contribute to cut graphene oxide into 3-5 nm GQDs in one step. The quantum yield of this method is 43%. Fluorescence lifetime of these blue GQDs is 1.67 ns. The Zigzag-type site and defect state of the triplet carbene radical lead to the excitation wavelength dependence of GQDs, and the optimal excitation and emission wavelengths are 330 nm and 400 nm, respectively. The boundary effect and amphiphilicity of quantum dots make GQDs possess abundant functional groups, vacancy defects and high dispersion, which results in GQDs exhibits good water solubility. RAW264.7 macrophages are incubated with different concentration in DEME medium for 24 h, 48 h and 72 h to evaluate cell. The survival rate of RAW264.7 cells is significantly dependent on the concentration and time of GQDs. CCK-8 and dead/alive staining show that GQDs have high biocompatibility. The effect of 200 mg/L GQDs on apoptosis of RAW264.7 cells is revealed by the scatter plot of bivariate flow cytometry. Under the stimulation of LPS+INF-γ, the expression of TNF-α was increased in RAW264.7 cells, which co-acted with other cytokines to participate in the immune response of RAW264.7 cells in vitro, and mediated the production of IL-1β inflammatory factor in RAW264.7 cells, thereby inducing apoptosis of RAW264.7 cells. The results of RT-qPCR showed that GQDs can inhibit the growth of RAW264.7 cells in vitro, and stimulate them to increase TNF-α expression in RAW264.7 cells, which make cell membrane rupture and produce IL-1β inflammatory factors to induce cell apoptosis. The high biocompatibility of GQDs is attributed to the rich oxygen-containing functional groups (―COOH, ―OH, and CO) on the surface of GQDs, which makes its surface negatively charged and easy to be swallowed into the cell interior when interacting with the cell membrane with low affinity. Transmission electron microscopy (TEM) observed that the GQDs were swallowed into the cells. Furthermore, laser confocal results displayed that blue GQDs has certain ability of cell imaging in vitro.Conclusion The water solubility, low toxicity, fluorescence properties and the induction effect of inflammatory factors of GQDs provide broad prospects for their application in the field of immunotherapy and cell imaging in the future.

Abstract:Objective To explore the mechanism of treadmill exercise against type 2 diabetes mellitus (T2DM) with non-alcoholic fatty liver disease (NAFLD) based on the regulator effects of exercise on ferroptosis.Methods Eight 8-week-old male m/m mice were used as control group (Con, n=8), and db/db mice of the matched age were randomly divided into T2DM model group (db/db, n=8), exercise group (db+Exe, n=8), p38 mitogen-activated protein kinase (MAPK) inhibitor group (db+SB203580, n=8) and exercise combined with p38 MAPK inhibitor group (db+Exe+SB203580, n=8). After one-week adaptive feeding, the mice in the db+Exe group and db+Exe+SB203580 group underwent moderate intensity treadmill exercise for 40 min/d, 5 d/week lasting 8 weeks. The db+SB203580 group and db+Exe+SB203580 group were treated with SB203580 (a specific inhibitor of p38 MAPK) with a dose of 5 mg/kg, 5 d/week for 8 weeks. And the exercise intervention was performed 2 h later after the intraperitoneal injection of SB203580. The body weight and fasting blood glucose of mice were measured regularly every week during the experiment. After 24 h of the last intervention, the mice were weighted, the liver tissues were taken, weighted and the liver index was calculated. The pathological changes of liver were determined by Oil Red O and hematoxylin-eosin (HE) staining. The levels of blood lipids, liver function, Fe²⁺ and oxidative stress markers of liver were measured by enzyme linked immunosorbent assay (ELISA). The related mRNA expression levels of lipogenesis and inflammation were evaluated by quantitative reverse transcriptase-mediated PCR (qRT-PCR). The related protein expression levels of lipogenesis and ferroptosis in liver were determined by immunohistochemical (IHC) staining and Western blot.Results The body weight, fasting blood glucose, liver index, blood lipid and transaminase levels in the db/db group were significantly increased compared with the Con group. HE and Oil Red O staining showed severe lipid accumulation and ballooning change in the liver of db/db mice. Biochemical tests showed that Fe²⁺ and MDA level of liver constitution homogenate increased, while GSH level decreased significantly. The results of qRT-PCR showed that the mRNA levels of MCP-1, IL-6, SREBF1 and ACC1 in liver tissue of db/db mice were all significantly increased. Western blot results showed that the expression levels of SREBF1, ACC1 increased, ferroptosis relative proteins were significantly decreased. The 8 weeks of exercise significantly reduced the rise in body weight, blood glucose, liver index and blood lipid levels in db/db mice. Exercise intervention also alleviated hepatic steatosis and reduced the expression levels of Fe²⁺, MDA, MCP-1, IL-6, ACC1 and SREBF1, upregulated the expression levels of GSH, NRF2, HO-1, SLC7A11 and GPX4 in liver tissue of db/db mice. The intervention of exercise combined with SB203580 significantly down-regulated the mRNA expression levels of ACC1, MCP-1, IL-6, reduced the levels of Fe²⁺ and MDA, and up-regulated the level of GSH in db/db mice. Compared with the db+Exe group, the expression of Fe²⁺, MDA, MCP-1, and SREBF1 in the liver of the db+Exe+SB203580 group mice significantly increased, while the expression level of GSH and expression levels of ferroptosis relative proteins also significantly decreased. In addition, compared with db+SB203580 group, the iron accumulation and lipid peroxidation in the liver of db+Exe+SB203580 group were significantly improved.Conclusion The 8-week treadmill exercise can effectively alleviate liver injury and steatosis, and its mechanism may be related to the inhibition of hepatocyte ferroptosis through p38 MAPK signal.

Abstract:Objective Asthma is a common chronic inflammatory airway disease, and severe asthma poses a significant challenge in its diagnosis and management. Immune cells are involved in and altered by asthma pathogenesis, and several lipid metabolites can serve as diagnostic markers for the disease. In this study, we investigated the characterization of severe asthma at the metabolic and cellular level.Methods Differential metabolites in blood samples from severe asthma (41 cases) and controls (18 cases) were screened using multifactorial statistical analysis and independent samples t-tests; key pathways were identified by KEGG enrichment analysis, and biomarkers were characterized based on ROC curves; immune cell types and proportions in the blood were identified based on the results of cell-type annotations (5 severe and 3 control cases); and single-sample Gene Enrichment Analysis (ssGSEA) to investigate the characterization of differential metabolic pathways in single cells.Results Compared with controls, the abundance of 28 metabolites was increased and the abundance of 13 metabolites was decreased in the blood of patients with severe asthma (P<0.05); the differential metabolites were enriched in 4 pathways: sphingolipid metabolism, glycerophospholipid metabolism, nicotinate and nicotinamide metabolism, and histidine metabolism. Among them, 13 differential metabolites could be used as biomarkers for the diagnosis of severe asthma, including L-glutamic acid (AUC=0.809), nicotinamide (AUC=0.886), phytosphingosine (AUC=0.882), and sphinganine (AUC=0.893). In single-cell transcriptome analysis, 5 key cell types were identified: CD4⁺ T cells, CD8⁺ T cells, NK cells, B cells, and monocytes. The number of NK cells was increased in patients with severe asthma, and severe asthma exhibited more frequent cellular communication, particularly dense communication between CD8⁺ T cells and other cell types. In healthy samples, monocytes were the primary cells engaging in dense communication. Single-sample gene enrichment analysis (ssGSEA) showed that 4 pathways enriched for differential metabolites had lower scores (P<0.01) in CD4⁺ T and CD8⁺ T cells in severe patients, and it was hypothesized that the expression of genes associated with these pathways was suppressed in these two types of cells. The suppressed genes included DGKA and NT5C3A, which are associated with immune processes. We observed that these genes play key roles in the regulation of T cell signaling, activation, differentiation, and immune responses.Conclusion L-glutamic acid, nicotinamide, phytosphingosine, and sphinganine can be used as biomarkers for the diagnosis of severe asthma; genes of the severe asthma-associated pathway were suppressed in CD4⁺ T cells and CD8⁺ T cells.

Abstract:Objective Coronavirus is a class of long-standing pathogens, which are enveloped single-stranded positive-sense RNA viruses. The genome all encodes 4 structural proteins: spike protein (S), nucleocapsid protein (N), membrane protein (M), and envelope protein (E). The nucleocapsid protein (NP) serves as a key structural component of coronaviruses, playing a vital function in the viral life cycle. NP acts as an RNA-binding protein, with a critical role in identifying specific sequences within the viral genome RNA, facilitating the formation of ribonucleoprotein (RNP) complexes with viral RNA to stabilize the viral genome and contribute to viral particles assembly. The NP consists of two primary structural domains, the N-terminal domain (NTD) and the C-terminal domain (CTD). The NTD is primarily responsible for RNA binding, whereas the CTD is involved in polymerization. The N protein demonstrated to trigger the host immune response and to modulate the cell cycle of infected cells by interacting with host proteins. The NP, one of the most abundant protein in coronaviruses, is essential in understanding the pathogenic mechanism of coronaviruses through its interaction with host factors, which response for determining the virus pathogenicity. HCoV-229E is a widely distributed coronavirus that typically causes mild upper respiratory tract diseases, accounting for a significant portion of common cold cases. However, its pathogenicity is notably lower compared to other coronaviruses like MERS-CoV, SARS-CoV, and SARS-CoV-2. The exact molecular mechanism behind remains unexplained, and how HCoV-229E N protein influences virus replication, host antiviral immunity, and pathogenesis need to be further explored.Methods Proximity labeling-mass spectrometry technique and bioinformatics analysis were used to screen for potential host factors interacting with the NP of human coronavirus 229E (HCoV-229E). In this study, a recombinant adenovirus Ad-V5-NP^HCoV-229E-TurboID was constructed to express the fusion protein of HCoV-229E NP and biotin ligase (TurboID). A549 cells were infected with the Ad-V5-NP^HCoV-229E-TurboID. After 30 min biotin treatment, NP interacting proteins were labeled with biotin by biotin ligase, and subsequently isolated with streptavidin cross-linked magnetic beads. The potential interacting proteins were identified using label-free proteomic mass spectrometry and further validated through immunoprecipitation and immunofluorescence assays.Results We identified a total of 584 potential interacting proteins. Gene ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis highlighted the enrichment of glycogen synthase kinase (GSK)3A and GSK3B in the glycolysis/gluconeogenesis pathway, indicating HCoV-229E NP connection to diabetes through aberrant activity. Moreover, SARS-CoV-2 infection can exacerbate hyperglycemia and metabolic dysregulation in diabetic individuals by activating the ACE2 receptor. Moreover, SARS-CoV-2 was observed to cause potentially harm to pancreatic β-cells and leading to insulin deficiency, which not only worsens the condition of diabetic patients but also raises the possibility of new-onset diabetes in non-diabetic individuals. We demonstrated that GSK3A and GSK3B interacted with NP of HCoV-229E, suggesting that the NP may engage in various coronavirus pathogenic processes by interacting with GSK3.Conclusion These findings suggest that proximity labeling-mass spectrometry technique is a valuable tool for identifying virus-host interaction factors, and lay the foundation for future investigations into the mechanisms underlying coronavirus replication, proliferation, and pathogenesis.

Abstract:Objective The detection of RNA single nucleotide polymorphism (SNP) is of great importance due to their association with protein expression related to various diseases and drug responses. At present, splintR ligase-assisted methods are important approaches for RNA direct detection, but its specificity will be limited when the fidelity of ligases is not ideal. The aim of this study was to create a method to improve the specificity of splintR ligase for RNA detection.Methods In this study, a dual-competitive-padlock-probe (DCPLP) assay without the need for additional enzymes or reactions is proposed to improve specificity of splintR ligase ligation. To verify the method, we employed dual competitive padlock probe-mediated rolling circle amplification (DCPLP-RCA) to genotype the CYP2C9 gene.Results The specificity was well improved through the competition and strand displacement of dual padlock probe, with an 83.26% reduction in nonspecific signal. By detecting synthetic RNA samples, the method demonstrated a dynamic detection range of 10 pmol/L-1 nmol/L. Furthermore, clinical samples were applied to the method to evaluate its performance, and the genotyping results were consistent with those obtained using the qPCR method.Conclusion This study has successfully established a highly specific direct RNA SNP detection method, and provided a novel avenue for accurate identification of various types of RNAs.

Abstract:Objective Glutathione (γ-glutamyl-L-cysteinylglycine, GSH) is the most abundant non-protein compound containing sulfhydryl (―SH) groups in cells. It serves as a source of reducing equivalents, effectively neutralizing harmful reactive substances, and playing a crucial role in maintaining cellular redox balance. Therefore, sensitive detection and accurate measurement of GSH levels in tissues are of great importance. In this work, we presents a novel method for GSH detection utilizing electron paramagnetic resonance (EPR) spectroscopy.Methods Initially, ABTS (2,2"-azino-bis(3-ethylbenzothiazoline-6-sulfonate acid)) solution was mixed with K₂S₂O₈ solution and reacted in the dark for 12 to 16 h to prepare ABTS·⁺ solution, which was then quantified using UV-Vis spectroscopy. Subsequently, the concentration of glutathione (GSH) was determined based on the changes in the EPR signal of ABTS·⁺. On this basis, the optimal reaction time and temperature were explored to establish a standard equation correlating the EPR signal intensity of ABTS·⁺ with GSH concentration. Finally, the derived standard curve was employed to quantitatively analyze the GSH concentration in whole blood from C57BL/6J mice, and the results were compared with those reported in the literature to verify the accuracy of the method.Results The experimental results demonstrate that this method has a linear detection range from 50 nmol/L to 15 μmol/L for GSH, spanning two orders of magnitude, with a limit of detection (LOD) at 0.50 nmol/L. The measured GSH content in mouse whole blood is (10 660±706) nmol/g Hb, which agrees with the value of (11 200±237) nmol/g Hb as previously reported. Furthermore, a similar method was developed for detection of glutathione disulfide (GSSG) at higher reaction temperature.Conclusion This article presents a novel assay for the rapid detection of GSH using the intensity of EPR signal from ABTS·⁺ as indicator. This method demonstrates enhanced detection sensitivity and a broader linear range compared to conventional colorimetric methods. Furthermore, we have extended the application of this method to detect GSH content in blood samples efficiently and accurately, offering valuable information for assessing tissue redox balance, thus holding significant potentials.

Abstract:As a rapidly developing frontier discipline, structural biology has penetrated into every field of life science research. The course of “Structural Biology” plays an important role in expanding the knowledge system of undergraduate students and promoting students’ scientific spirit and innovation. For the high-quality training of highly skilled talents, we aimed to promote the original innovation of students, the ability of thinking, and the ability of engineering practice. The trinity education concept, including shape of the value, passing on knowledge, and ability cultivation, was applied. During the reform, we explored a step-by-step course content and searched for factors involved in ideological and political education. Based on the problem-based learning (PBL) method, a hybrid teaching model was designed to cultivate the problem-thinking and problem-solving skills of students. Meanwhile, a number of evaluation systems for students and teachers were established, which may be generally adopted for the course of “Structural Biology”. The survey data suggested that the exploration has a good effect on teaching and training and is conducive to the cultivation of research-oriented, comprehensive, innovative talents under the background of “New Engineering”.