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    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 0-0.
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    • CJBMB: 40 Years of Biochemistry and Molecular Biology in China
  • An Exploratory Experiment on the Dynamic Structural Change of ATP Synthase
    LIU Yi-Xuan, LIU Yang, ZHU Wen-Yuan, HU Xiao-Qian, CHANG Zeng-Yi, QIN Yong-Mei, WANG Qing-Song
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 625-631. https://doi.org/10.13865/j.cnki.cjbmb.2025.02.1467
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    The lab module of exploratory experiment is newly designed in the practical course of biochemistry. Here we describe one of the experimental projects, and it originates from new scientific research results on the dynamic structure of ATP synthase. This exploratory experiment is organized in the form of real scientific research, which would fully mobilize the initiative and creativity of students in learning theoretical knowledge and experimental technology. Students work in groups and start with reference reading. Through cooperation, they must develop certain experimental plan, handle samples with photocrosslinking technique and utilize the high-throughput electrophoresis method to analyze the dynamic structural change of ε subunit in ATP synthase under different physiological conditions. High quality results from high-throughput electrophoresis can only be obtained through optimized operation and treatment, from which students would experience the process of technological innovation. The teaching process of this lab module embodies the student-centered teaching concept and is widely approved and supported by students. The project of ATP synthase closely combines the content of lab course with cutting-edge technology. Students can deeply experience the importance of experimental technology innovation in solving scientific problems. The practical ability of students would be comprehensively improved through this lab module.
    • Reviews
  • The Role and Mechanism of m6A (N6-methyladenosine) and Its Regulatory Proteins in the Occurrence and Development of Infectious Diseases
    ZHANG Yan-Yan, WU Min, LIU Lin
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 632-644. https://doi.org/10.13865/j.cnki.cjbmb.2025.03.1414
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    N6-methyladenosine (m6A) is one of the most common post-transcriptional modifications in eukaryotic RNA, which has a profound impact on the structure, fate and function of RNA. The related proteins that regulate RNA m6A modification mainly include three categories: methyltransferase (writers), demethylase (erasers), and reading proteins (readers). The discovery of these enzymes suggests that m6A modification is a dynamic and reversible process that can influence RNA stability, translation efficiency, and localization by regulating RNA structure. Changes in the structure of RNA directly determine the fate, function and metabolism of RNA, and then regulate cell proliferation, differentiation and apoptosis, and participate in the occurrence and development of a variety of diseases including metabolic diseases and cancer. In recent years, increasing studies have revealed the important role of m6A modification in regulating innate immune system homeostasis. The immune system is crucial for defending against infection by bacteria, viruses and other pathogens, and m6A modification regulates the activity and response of immune cells by affecting the expression and function of immune-related genes. This review summarizes recent reports on the biological significance of m6A modifications, the major regulatory functions of protein, and how they work together to maintain the homeostasis of the immune system. How m6A modifications affect the expression and signaling of pattern recognition receptors (PRRs) and its role in antiviral and antimicrobial immune responses are summarized. This review also analyzes the effects of m6A modification on the differentiation, activation and function of T and B cells, especially in vaccination and autoimmune diseases. Furthermore, the regulatory mechanisms of m6A modification in different infectious diseases, such as viral, bacterial, and parasitic infections, and its potential as a potential therapeutic target are also reviewed. The problems and challenges in current research and future research directions are presented, including the development of new technology and therapeutic approaches to better understand and utilize the m6A modification in immune regulation. Combined with our own research results, the role of m6A and its regulatory proteins in regulating immune system function and immune response in infectious diseases is elaborated. By synthesizing the existing literature and our findings, this review aims to provide researchers with a comprehensive and detailed perspective on the recent advances in m6A and its regulatory proteins in regulating immune system function and immune responses in infectious diseases, laying the foundation for further research and clinical applications.
  • Mechanism of Lactylation in Ischemic Heart Diseases
    WANG Peng-Fei, HU Feng-Li, GU Guo-Qiang
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 645-652. https://doi.org/10.13865/j.cnki.cjbmb.2025.03.1468
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    Ischemic heart disease (IHD) is one of the major threats to global health, characterized by complex and incompletely elucidated pathogenesis. Recently, with the continuous advancement of epigenetic research, lactylation (Kla), a newly discovered type of protein post-translational modification, has gradually attracted attention. Kla significantly influences the pathophysiological processes and cellular molecular functions of IHD by directly affecting gene transcription, signal transduction, and metabolic pathways. Kla extensively occurs in both histones and non-histone proteins and participates in regulating protein functions involved in various pathological processes. By modulating enzymatic activities and signal transduction pathways, Kla affects multiple processes in cardiomyocytes, including energy metabolism, inflammatory response, angiogenesis, lipid metabolic disorders, apoptosis, fibrosis, and myocardial repair. Although current studies on specific mechanisms and therapeutic targets of Kla in IHD remain limited, its potential therapeutic value cannot be overlooked. This review summarizes the mechanisms and research progress of Kla in critical pathological stages of IHD, such as myocardial infarction, myocardial ischemia-reperfusion injury, heart failure, and cardiac hypertrophy. Furthermore, we discuss the potential therapeutic targets and application prospects of Kla, aiming to provide insights and directions for identifying effective intervention strategies and opening new avenues for the prevention and treatment of IHD.
  • Development and Application of the PROTAC Technique in Castrate-resistant Prostate Cancer
    WANG Si-Jia, XU Cheng, CHEN Wen-Min
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 653-663. https://doi.org/10.13865/j.cnki.cjbmb.2025.03.1314
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    Abnormal signaling in the androgen receptor (AR) signaling pathway is critical for prostate cancer development and progression, so inhibition of AR activity through androgen deprivation therapy (ADT) is an important means to control the development of prostate cancer in the early stage. However, most patients relapse and develop castrate-resistant prostate cancer (CRPC) within 6~20 months. Surgery and radiotherapy are still the major treatments for CRPC, but there are adverse effects such as urinary symptoms and sexual dysfunction. The first and second generatiosn of novel AR inhibitors can effectively treat CRPC. However, resistance to these chemicals is inevitable, and thus many patients may experience recurrence. Resistance to AR inhibitors mainly consists of AR mutations, splice variant formation and amplification, which have been shown to play an important role in CRPC. Also, aberrant activation of cyclin dependent kinase (CDKs) and epigenetic alterations (e.g. histone modifications and DNA methylation) have been reported to be associated with prostate cancer progression. Proteolysis targeting chimeras (PROTACs) have unique advantages in CRPC therapy by virtue of their unique mechanism of action, ability to target non-druggable proteins, and specific binding to targets. In this review, we summarize the development of PROTAC technology for the treatment of CRPC by targeting different structural domains of AR, CDKs and epigenetic markers, and discuss the future prospects and challenges of PROTACs in the therapeutic field.
  • Epigenetic Regulation and Treatment of HIV Infection: Advances and Challenges
    DUAN Jiang-Kai, CHEN Liang
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 664-677. https://doi.org/10.13865/j.cnki.cjbmb.2024.03.1236
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    Currently, acquired immune deficiency syndrome (AIDS) has emerged as a global public health crisis that profoundly compromises human immune defenses. By systematically dismantling the immune system, human immunodeficiency virus (HIV) renders individuals vulnerable to opportunistic infections and malignancies, ultimately culminating in AIDS progression. It is urgent to eradicate the latent HIV virus and achieve a functional cure, thus limiting the development of AIDS and improving the quality of patients. Epigenetics investigates heritable alterations in gene expression that occur independently of DNA sequence. The intricate regulation of HIV gene expression is orchestrated through multifaceted epigenetic mechanisms involving both viral and host factors. Understanding the epigenetic mechanisms associated with HIV infection is crucial for clearing latent viruses and achieving control and treatment of AIDS in the future. Therefore, we will discuss the epigenetic regulatory patterns and mechanisms involved in HIV infection, particularly emphasize on four principal mechanisms: DNA methylation, histone modification, non-coding RNA regulation and RNA modification. We comprehensively analyze how these regulatory factors influence the viral life cycle, particularly regarding latency establishment, reactivation dynamics, and persistent infection maintenance. Furthermore, we delineate the interplay between epigenetic regulators and key cellular signaling pathways during HIV pathogenesis. The review culminates in a critical appraisal of recent breakthroughs and persistent challenges in epigenetics-based therapeutic strategies, while highlighting innovative approaches for functional cure development. By elucidating the pivotal role of epigenetic regulation in HIV latency, this review aims to establish a novel theoretical foundation and innovative research directions for next-generation AIDS therapeutics rooted in epigenetic modification.
  • Biological Characteristics of Cardiolipin in Cardiovascular Diseases
    ZHANG Hui-Fei, JIANG Yue-Hua
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 678-686. https://doi.org/10.13865/j.cnki.cjbmb.2025.03.1325
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    Cardiolipin (CL) is a special type of polyglycerophospholipid, primarily synthesized in the mitochondrial inner membrane and cristae, and serves as a key component for mitochondrial function. It plays an essential role in the cellular membrane, mitochondrial inner membrane and energy metabolism, especially in maintaining the stability of oxidative phosphorylation and the electron transport chain. Abnormal metabolism of cardiolipin is closely associated with the occurrence of various cardiovascular diseases, particularly in genetic disorders such as Barth syndrome (BTHS). Moreover, the role of cardiolipin peroxides in cardiovascular diseases has been increasingly recognized. Studies have shown that cardiolipin peroxidation not only leads to damage of the mitochondrial inner membrane but also promotes the generation of reactive oxygen species (ROS), thereby enhancing oxidative stress within the cell. Abnormal metabolism of cardiolipin is also closely related to the pathogenesis of atherosclerosis, diabetic cardiomyopathy, hypertension, and other diseases. Regulating cardiolipin metabolism and repairing its functional defects may offer potential strategies for treating these diseases. This review discusses the synthesis, degradation, and remodeling processes of cardiolipin, and explores its significant role in cardiovascular diseases. The synthesis of cardiolipin relies on various enzymes within the mitochondria, while its remodeling involves key enzymes such as phosphatidyltransferases. Abnormal metabolism of cardiolipin, particularly the CL remodeling defects caused by tafazzin gene mutations in BTHS patients, leads to mitochondrial dysfunction, reduced ATP synthesis, increased oxidative stress, and ultimately results in myocardial and other tissue damage.
  • Neuroprotective Mechanism of Exercise on Alzheimer’s Disease: Role of Oxidative Stress
    ZENG Fan-Xi, ZHAO Ren-Qing, WANG Bin
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 687-695. https://doi.org/10.13865/j.cnki.cjbmb.2025.02.1403
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    Alzheimer’s disease (AD) is a neurodegenerative disorder primarily affecting memory, learning, and cognitive functions. It poses a significant health concern for the elderly, but effective treatments are lacking. Its main pathological features are amyloid β (Aβ) deposits forming senile plaques (SPs) and neurofibrillary tangles (NFTs) formed by hyperphosphorylated tau (p-Tau). These pathological changes often induce oxidative stress, which is an important pathological mechanism in AD. Oxidative stress is closely associated with Aβ and Tau deposition and is a potential target for intervention in the treatment of AD. However, the pathological mechanisms leading to AD are multifactorial, and AD oxidative stress often interacts with other mechanisms to jointly influence the AD process. Therefore, this paper focuses on the regulatory relationship between mitophagy, neuroinflammation, neuronal apoptosis and nuclear factor erythroid 2-related factor 2 (Nrf2) and oxidative stress. By elucidating the relationship between the pathological features, oxidative stress and its regulatory mechanism of AD, potential effective intervention targets were found. At present, numerous studies have indicated that exercise can alleviate oxidative stress in AD and improve cognitive function, but the underlying molecular mechanisms require further clarification. Therefore, we further discussed the mechanism by which exercise regulates oxidative stress and related molecular signaling pathways, and clarified that exercise may ameliorate AD oxidative stress by affecting these signaling pathways, thereby improving AD-related pathological features and cognitive function. It is helpful to understand the pathogenesis of AD from the perspective of molecular mechanism and provide theoretical support for scientific and effective exercise intervention to prevent and cure AD.
    • Research Papers
  • miR-29-TET2 Inhibits Lipid Accumulation in Hepatocytes by Activating the Autophagy Pathway
    SHEN Rui-Li, LI Han-Bing, FAN Yu-Wei, CHENG Ni-Hong, WU Wen-Jing, ZHANG Jin
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 696-706. https://doi.org/10.13865/j.cnki.cjbmb.2025.03.1021
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    The incidence of non-alcoholic fatty liver disease (NAFLD) has been increasing annually. Current primary treatment strategies involve dietary modifications and increased physical activity to alleviate symptoms, yet there is a notable lack of targeted pharmacological interventions. Members of the micro RNA-29 (miR-29) family (miR-29a, miR-29b, miR-29c) are known to play a critical regulatory role in lipid metabolism within hepatocytes; however, the underlying mechanisms remain to be elucidated. This study aims to identify the target genes and associated signaling pathways of the miR-29 family, thereby providing potential therapeutic targets for the development of NAFLD treatments. Firstly, the human liver cell line HepG2 was utilized as a model for adipogenic induction, and miR-29a/b/c-3p mimics were individually transfected. Through methods such as Oil Red O staining and triglyceride (TG) quantification, it was observed that the miR-29 family members significantly inhibited lipid accumulation in hepatocytes (P<0.05). Subsequently, qRT-PCR and Western blot were utilized to detect the expression levels of adipogenic marker genes (fatty acid synthase (FAS), acetyl coa carboxylase (ACACA) , stearoyl-coenzyme a desaturase1 (Scd1)) and autophagy marker genes (sequestosome 1 (SQSTM1, also known as p62), autophagy related gene 5 (Atg5)), and the results indicated that the members of the miR-29 family could significantly suppress the expression of FAS, ACACA, Scd1, and p62 genes in hepatocytes, while significantly enhancing the level of the Atg5 gene. Further investigations using signaling pathway activity analysis and dual luciferase reporter assays confirmed that the miR-29a/b/c could suppress the mTOR signaling pathway activity and directly interact with the ten-eleven translocation 2 (TET2) gene. Finally, co-transfection experiments were performed to examine the potential synergistic effects among the miR-29-3p family members, and the results demonstrated that co-transfection of miR-29 family members more effectively inhibited lipid droplet accumulation in HepG2 cells and further suppressed the expression of the target gene TET2 compared to individual transfection. In summary, the miR-29 family members may reduce lipid accumulation in hepatocytes by inhibiting the mTOR signaling pathway via the TET2 gene, and they exhibit a positive synergistic effect.
  • The Role of Zinc Finger Structure Transcription Factors ZNF148 and SP5 on P53 Transcriptional Activity
    WANG Dai-Wei, ZHOU Chen, ZHANG Pin-Zheng, WANG Xu-Ying, LI Jia-Wen, MA Yu-Kai, YAN Jia-Qi, WANG Zhi-Ting, WANG Jia-Qi, GUO Zhi-Yi
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 707-715. https://doi.org/10.13865/j.cnki.cjbmb.2025.03.1514
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    P53 is a key tumor suppressor gene, which is regulated in many ways. Zinc finger 148 (ZNF148) and SP5, as zinc finger transcription factors (TFs), play important roles in tumor suppression and carcinogenesis. The regulatory relationship between these two TFs and p53 has not been reported. In this paper, Ishikawa and A549 cell lines with different p53 expression levels were used as research models to explore the transcriptional regulation of the P53 gene by ZNF148 and SP5. The data showed that there were differences in the expression of ZNF148 and SP5 in the two cell lines. The mRNA expression of ZNF148 in Ishikawa was 1.9 times higher than that of A549, and the mRNA expression of SP5 in A549 was 802.4 times that of ZNF148. Data showed that in Ishikawa cells, the expression of P53 decreased (81.8%) after ZNF148 knockdown, and increased (2.6 times) after SP5 overexpression. Transfection of si-SP5 and ZNF148 expression plasmids into A549 cells increased the mRNA expression of P53 by 6.6 times and 14.6 times, respectively. These results indicate that ZNF148 could activate, whereas SP5 could inhibit, P53 expression. The conserved cis-element of ZNF148 and SP5 TFs was found in the region of the P53 promoter by bioinformatics methods. The data from dual luciferase reporter gene assay showed that the luciferase activity of ZNF148 in Ishikawa and A549 cells was increased by 2.1-fold and 4.2-fold compared with the control group (P<0.05). Compared with the control group, the normalized relative luciferase activity of transfected SP5 decreased by 77.1% and 35.7% (P<0.05). However, when the cis-element of ZNF148 and SP5 was mutated, the effect disappeared. Further transfection of ZNF148 and SP5 with different ratios revealed that SP5 could reverse the transcriptional activation of P53 by ZNF148. Studies have shown that ZNF148 shares a common site with SP5, and the ratio of the two TFs may influence the transcriptional activity of P53. The expression of the Wnt pathway and the cell proliferation rate after knockdown of ZNF148 and SP5 were further studied to explore the role of the two TFs. Our data show that ZNF148 and SP5 could regulate the transcriptional activity of P53, and their expression levels and interaction may be the key factors regulating P53 expression.
  • Identification and Expression Analysis of The Glycyrrhiza SQE Gene Family Members under Abiotic Stress
    LI Yue-Tao, CHENG Lin-Yuan, YAO-Hua, SHEN Hai-Tao
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 716-729. https://doi.org/10.13865/j.cnki.cjbmb.2025.05.1525
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    Glycyrrhizic acid is one of the key bioactive components in licorice, known for its liver-protective and antiviral effects. Squalene epoxidase (SQE) is a crucial enzyme in the biosynthetic pathway of glycyrrhizic acid. However, there is limited research on the systematic analysis of the SQE gene family and its function in licorice. This study aims to explore the role of the SQE gene family in glycyrrhizic acid synthesis through bioinformatic analysis, expression specificity, and correlation with glycyrrhizic acid content. The results showed that the three medicinal species of licorice contained a total of 11 SQE genes. Among them, both Glycyrrhiza glabra and Glycyrrhiza inflata had four SQE genes, while Glycyrrhiza uralensis had three. Highly homologous SQE genes exhibited similar expression patterns and were located at similar chromosomal positions. Different SQE genes displayed distinct expression characteristics. Specifically, GgSQE1, GiSQE1, GuSQE1, and GuSQE3 were primarily expressed in the roots, while GgSQE3 was highly expressed in the whole plant of licorice. Under 15% PEG6000 and 150 mmol/L NaCl treatments at different time points during seedling stages of different licorice species, the expression patterns of GgSQE1, GgSQE3, GiSQE1, GiSQE3, and GuSQE1 exhibited trends similar to the changes in glycyrrhizic acid content. Further analysis revealed that the promoter regions of these genes contained multiple stress-responsive elements, suggesting that SQE1 and SQE3 may be involved in glycyrrhizic acid synthesis following abiotic stress in licorice. The findings of this study provide candidate genes for future breeding programs aimed at improving glycyrrhizic acid content and lay a foundation for further research into the molecular mechanisms by which abiotic stress enhances glycyrrhizic acid production.
  • Treating Type 2 Diabetic Nephropathy by Down-regulating NOX4 to Inhibit the Oxidative Stress Pathway in Mesenchymal Stem Cells
    FENG Shu-Qi, JIN Guo-Rong, XUE Qun-Hang, HE Min, WANG Ze-Hang, YAO Jia-Xin, CHEN Long, WANG Yu-Jiao, ZHANG An-Xiu, HE Sheng, ZHOU Bing-Rui, XIE Jun
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 730-740. https://doi.org/10.13865/j.cnki.cjbmb.2025.03.1425
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    Diabetic nephropathy (DN) is a serious complication of diabetes mellitus and a leading cause of end-stage renal diseases. In DN patients, key pathological mechanisms include proteinuria, glomerulosclerosis, and fibrosis, largely driven by poor glycemic control and oxidative stress caused by prolonged hyperglycemia. This stress damages renal podocytes and triggers inflammatory mesenchymal infiltration of renal tubular cells, exacerbating the progression of proteinuria and fibrosis. Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) offer promising potential for treating DN due to their strong anti-oxidative properties. In this study, we developed a DN mouse model and treated the mouse via tail vein injections of hUC-MSCs (1×106 cells/mouse). The results indicated that hUC-MSCs significantly lowered fasting blood glucose levels (22.5 ± 3.0 vs 14.7 ± 1.1, P < 0.01) and improved glucose tolerance, as shown by intraperitoneal glucose tolerance test (IPGTT) results (P < 0.05). Additionally, the renal function improved in hUC-MSCs-treated mice, with marked reductions in oxidative stress markers, including blood urea nitrogen (BUN), urinary creatinine (Ucr), urinary protein (PRO), superoxide dismutase (SOD), and malondialdehyde (MDA) (P < 0.05). Histological analyses through hematoxylin-eosin (H&E), Periodic Acid-Schiff (PAS), and Sirius red staining demonstrated alleviation of glomerular mesangial hyperplasia, glomerular hypertrophy, and tubular inflammation. Furthermore, hUC-MSCs treatment downregulated the expression of oxidative stress-related proteins, such as NADPH oxidase 4 (NOX4) and thioredoxin-interacting protein (TXNIP), and reduced reactive oxygen species (ROS) production (P < 0.05). Meanwhile, human renal cortical proximal tubule epithelial cells (HK-2 cells) were selected for validation in vitro experiments using high glucose treatment followed by supernatants of hUC-MSCs (MSC-CM), and Western blotting showed that the expression of both NOX4 and TXNIP was inhibited (P < 0.05) and ROS expression was reduced. In conclusion, hUC-MSC treatment effectively lowered blood glucose levels and improved renal function in DN mice, likely through the suppression of NOX4 expression and TXNIP-mediated oxidative stress.
  • Investigations into the Mechanism of Phycocyanin in Modulating the Wip1/p53 Pathway to Induce Apoptosis in Human Hepatocellular Carcinoma HepG2 Cells
    JIA Yun-Xi, HUO Da, YAO Chao, LI Min, LIU Fu-Ling, YUAN Hong, XUE Hui-Ting, HU Rui-Ping
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 741-752. https://doi.org/10.13865/j.cnki.cjbmb.2025.03.1475
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    Hepatocellular carcinoma (HCC) is difficult to detect in its early stages and current treatment methods are associated with significant side effects and a high risk of developing drug resistance. This study aims to investigate the effect of phycocyanin (PC) on the apoptosis of human HCC HepG2 cells and its potential mechanism. HepG2 cells were treated with PC at concentrations of 0.1, 0.25, 0.5, 1, 2.5, 5, and 10 μg/mL for 12 h, and with 10 μg/mL PC and 2.5 μmol/L Wip1 inhibitor (Wip1i) alone or in combination for 12 and 24 h, respectively. Cell proliferation levels were assessed using the CCK-8 cell proliferation-toxicity assay kit. Apoptosis levels were measured by Annexin V-FITC/Propidium Iodide double staining combined with flow cytometry. TMT (Tandem Mass Tag) proteomics quantitative technology was applied to analyze differential protein expression. Western blotting was used to detect the expression levels of Wip1, p53, and phosphorylated-p53 (Ser15) proteins. The CCK-8 assay revealed that PC effectively inhibited HepG2 cell proliferation in a concentration-dependent manner, with a half-maximal inhibitory concentration (IC50) of 19.37 μg/mL. Flow cytometry results showed that PC significantly induced apoptosis, with an apoptosis rate of 30.40%. Quantitative proteomics analysis indicated that PC induced activation of the p53 pathway. The CCK-8 assay showed that Wip1i enhanced the cytotoxic effect of PC on HepG2 cells. Western blotting confirmed that PC inhibited Wip1 expression, induced p53 protein phosphorylation, and promoted the expression of total p53 protein. Additionally, Wip1i further enhanced PC-mediated activation of the p53 pathway, increasing the expression of p53 and pP53 (S15). In conclusion, PC may induce apoptosis by inhibiting the activity of the p53 negative regulator Wip1, thereby promoting apoptosis through the Wip1/p53 pathway.
  • Icaritin Targets P53 to Regulate DNA Damage Repair and FOXO Signaling Pathways to Inhibit Glioma Cell Growth
    LUO Zhi-Qiong, WANG Zhuo-Yi, WANG Yong-Ping, CHEN Xiao-Zhong, YU Jia, CHENG Sha, ZAN Ning-Ning, SUN Bao-Fei, LUO Heng
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 753-763. https://doi.org/10.13865/j.cnki.cjbmb.2025.03.1347
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    Icaritin (ICT) is an 8-isopentenylflavonoid, which is the main effective component of the traditional Chinese medicine Epimedium. Previously, we found that Icaritin inhibits the growth of glioblastoma (GBM) cells. Herein we aim to study the in vivo anti-GBM effectiveness of Icaritin and explore its mechanism. The results of MTT assay, flow cytometry, comet assay and cellular immunofluorescence assay in vitro showed that ICT inhibited the proliferation of four kinds of GBM cells, U87, U251, U118 and A172, induced early apoptosis (P＜0.001) and late apoptosis (P＜0.05) in U87 cells, induced DNA damage in U87 cells, and blocked the growth of U87 cells at the G0/G1 phase (P＜0.0001) in a concentration-time-dependent manner. In vivo subcutaneous tumor transplantation tumor experiments showed that feeding 200 mg/kg (P < 0.01) and 400 mg/kg (P < 0.001) ICT had a significant inhibitory effect on the growth of GBM subcutaneous tumors, and had no significant toxic effects on heart, liver, spleen, lung and kidney tissues. The results of network pharmacological analysis, molecular docking and cellular thermodynamic experiments showed that there were 26 possible target proteins between ICT and GBM, among which the expression of p53 in GBM tissues was significantly (P＜0.001) higher than in normal tissues, and the binding energy of ICT and p53 was lower; cellular thermodynamic experiments verified that ICT significantly enriched the level of p53 in the living cells of GBM, which indicated that ICT could target p53. The expression of key proteins in the DNA damage repair and apoptosis-associated FOXO signaling pathway was detected by ICT. The results showed that the expression of ATR (P < 0.01), P53 (P < 0.001), P21 (P < 0.05) and γ-H2AX (P＜0.05) was up-regulated, whereas the expression of Cyclin E1 (P < 0.01), E2F1 (P < 0.05), CDK2 (P < 0.01), Rb (P < 0.001), p-Rb (P < 0.0001) and WRN (P＜0.0001) expression were down-regulated. There was no significant change in the expression of FOXO1 in the FOXO pathway or a significant down-regulation of its phosphorylation level. This study demonstrated that ICT could effectively inhibit the growth of GBM cells in vivo. It targets p53 to regulate the DNA damage repair pathway and FOXO signaling pathway to induce GBM cell cycle arrest and apoptosis.
    • Techniques and Methods
  • Screening and Identification of Nanobodies Against β-Conglycinin
    CHANG Jia-Shu, SUN Hua-Bo, WANG Yu-Ting, WANG Xiao-Hui, YANG Bo, LIU Hong-Rui, LI Yue-Xin, SUN Yuan-Zhao, GU Shao-Peng, HE Jin-Xin
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 764-770. https://doi.org/10.13865/j.cnki.cjbmb.2025.03.1462
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    Soy is a vital source of plant carbohydrates. However, it poses significant allergenic risks, particularly to young children and animals. Among the various proteins in soy, β-conglycinin, which constitutes approximately 30% of total soy carbohydrates, is a primary allergen. Undigested β-conglycinin can lead to intestinal damage by inhibiting cell growth, disrupting the cytoskeleton, and inducing apoptosis. It can also enter the lymphatic and circulatory systems, triggering allergic reactions. Conventional ELISA methods for detecting β-conglycinin rely on polyclonal or monoclonal antibodies, which are limited by their large molecular weight, difficulty in accessing the protein core, and sensitivity to acidic and basic conditions. To address these limitations, this study aimed to develop nanobodies (Nbs) against β-conglycinin. Nbs, derived from the variable regions of heavy-chain antibodies found in camelids, have a molecular weight approximately one-tenth that of conventional antibodies. They offer advantages such as small size, stable structure, high specificity, and strong affinity. A female alpacas was immunized five times using β-conglycinin, which showed a heavy chain antibody potency of 1∶16 000 by ELISA. Peripheral blood lymphocytes were subsequently isolated and total RNA was extracted. The variable region of the heavy-chain antibody was amplified via PCR, and recombinant plasmids were constructed and transformed into the E. coli competency strain ER2738. The resulting library contained about 3.5×108 CFU/mL, which increased to 1.15×1012 PFU/mL after phage rescue, with a 100% Nbs gene insertion rate, indicating high diversity. Its Nbs phage output was significantly enriched by four rounds of solid-phase elution with an enrichment rate of 155.9. Four rounds of solid-phase panning yielded 35 positive clones, all of which shared the same amino acid sequence upon sequencing. The selected Nb was expressed in a prokaryotic system, and its binding ability to β-conglycinin was confirmed using Western blotting and ELISA. The results demonstrated excellent specificity and affinity. This research lays the groundwork for developing a rapid and efficient detection method for β-conglycinin using Nbs, potentially enhancing food safety and allergen management.
    • Cover Image Introduction
  • The three members of the miR -29 family- -miR- 29a , b, and c-function cooperatively ，akin to Nezha's“three heads and six arms," to enhance hepatocyte autophagy and mitigate non-alcoholic fatty liver disease
    Cover picture designer ZHANG Jin, WU Wen-Jing, SHEN Lu
    Chinese Journal of Biochemistry and Molecular Biology. 2025, 41(5): 771-771.
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