Objective: This study aimed to investigate the role of the LINE-1 retrotransposon-encoded ORF-1p protein in regulating the proliferation and morphological dynamics of 293T cells. Methods: RNAi technology was used to construct two 293T cell lines with down-regulated expression of LINE-1 ORF-1p. The interference efficiency was verified by qPCR and Western blot analysis. The expression and localization of ORF-1p were detected by immunofluorescence, the cell proliferation ability was detected by cell counting and clone formation experiments, and the changes in cell biological characteristics were detected by phalloidin and DAPI staining analysis. Results: Two 293T cell lines with LINE-1 knockdown were successfully constructed. The localization of ORF-1p expression was detected via immunofluorescence analysis, which revealed its predominant cytoplasmic distribution. Significant reduction in ORF-1p expression was detected following LINE-1 knockdown. Through cell counting and colony formation experiments, it was found that the proliferation ability and colony formation ability of cells were significantly reduced after ORF-1p was down-regulated (P< 0.05). The cytoskeleton was labeled with phalloidin and the nucleus was labeled with DAPI. Phase contrast microscopy and confocal microscopy were used to observe cell morphology. The results showed that the area of shL1-749 cells decreased and their roundness increased after ORF-1p was down-regulated (P< 0.05). Conclusion: Two LINE-1 knockdown cell lines were successfully established. It was found that down-regulating the ORF-1p protein can inhibit the growth of 293T cells and change their morphology, laying the foundation for further understanding of the molecular mechanism of LINE-1 in cell proliferation and regulation.
Objective: The polarization of fat tissue macrophages towards the M2 phenotype can induce the beige/browning of adipocytes through uncoupling protein 1 (UCP1)-dependent thermogenesis, thereby promoting energy expenditure in the body. The purpose of this study is to elucidate the regulatory role of cathepsin L (CTSL), a lysosomal cysteine protease, in brown adipogenesis and energy metabolism, focusing particularly on its effects on inflammatory phenotypes of adipose tissue macrophages under CTSL-deficient conditions. Methods: Systemic Cathepsin L-deficient (Ctsl-/-) mice and relevant cell models were utilized. Real-time fluorescence quantitative PCR and Western blot assays were employed to analyze the degree of browning in white adipose tissue (WAT) of mice. Flow cytometry was utilized to monitor the effect of Ctsl-/- on the M2 polarization of bone marrow-derived macrophages (BMDMs) in mice. Results: The results showed that Ctsl deletion did not affect body weight, but it did significantly increase body temperature and energy expenditure in mice. Concurrently, the protein levels of UCP1 increased in white adipose tissue (WAT), along with elevated mRNA expression of various browning markers, including Ucp1, Prdm16, Pgc1α, Cited1, Elovl3, and Ndufv2. Therefore, Ctsl-/- promoted beige/browning of WAT, leading to increased energy expenditure in the body. Further mechanistic studies revealed that Ctsl-/- enhanced M2 polarization of BMDMs, upregulating the mRNA expression of anti-inflammatory cytokines such as Il-4, Il-10, Il-13, Mrc1, and Mrc2. Conclusion: These findings indicate that Ctsl-/- promotes beige/browning of WAT by influencing the polarization of adipose tissue macrophages towards the M2 phenotype, thereby increasing energy expenditure. This provides a new target for the prevention and treatment of obesity and related metabolic disorders.
Objective: This study aims to isolate and identify avian pathogenic Escherichia coli (APEC) phages, and to investigate their biological characteristics, whole-genome sequences, and potential applications. Methods: Using the O78 serotype strain O78-1 as the host bacterium, a lytic phage was isolated from duck fecal samples via the double-layer agar method. Its morphology and size were observed by transmission electron microscopy. Biological characteristics such as the optimal multiplicity of infection (MOI), one-step growth curve, thermal stability, and pH stability were determined, and its therapeutic efficacy in vivo was evaluated. Results: A novel Dhakavirus genus myovirus, designated as PJNE213, was successfully isolated. Its complete genome is 170 713 bp long, with a GC content of 40%. It lacks known virulence genes, antibiotic resistance genes, and lysogenic genes. The phage PJNE213 exhibited lytic activity against the common APEC serotypes O1, O2, and O78, yet it did not lyse the probiotic E. coli Nissle 1917. The latent period of PJNE213 was 5 min, with a burst size of 140 PFU per cell. It maintained stable biological activity at 50℃ and within a pH range of 3 to 11. When used as a prophylactic agent, PJNE213 was shown to increase the survival rate of the Galleria mellonella model by 50%, while also prolonging and maintaining the survival time and body weight of chicks. Conclusion: The bacteriophage PJNE213 exhibits excellent safety, efficacy, and stability, making it a promising candidate as a reliable biological agent for the prevention and treatment of APEC.
Objective: This study aimed to obtain a novel potato I-type chymotrypsin inhibitor (buckwheat chymotrypsin inhibitor, BCI) with improved stability through the molecular modification of a buckwheat trypsin inhibitor, laying the foundation for the development of protease adsorbent ligands and other related applications. Methods: BCI was prepared by introducing a mutation to the key active site of the buckwheat trypsin inhibitor through site-directed mutagenesis technology. High-purity BCI was subsequently obtained by transformation and optimization of the induction conditions, and affinity chromatography purification. The inhibitory activity of BCI against chymotrypsin was determined, and its tolerance to heat, acidic conditions, and alkaline conditions was further analyzed. Results: The findings confirmed the successful construction of the pQE30-BCI mutant, with the optimal induction parameters at a final IPTG concentration of 0.4 mmol/L and an induction period of 3.5 h. Using a Ni2+-NTA magnetic agarose beads, 18.3 mg of electrophoresis-pure BCI were obtained from 1 L of culture medium. The semi-inhibitory concentration of BCI against chymotrypsin was 2.81 μmol/L with BTEE as the substrate. BCI exhibited remarkable thermal and pH stability, with no significant changes in its inhibitory activity after it was exposed to boiling water for 1 h or treated at pH levels ranging from 2.0 to 12.0 for 1 h. A concentration of 10 mmol/L of Mg2+, H2O2, and KSCN did not significantly affect the activity of BCI. However, Zn2+, dimethylacetamide and isopropanol reduced the activity of BCI. The results of fluorescence spectroscopy analysis showed that Zn2+ can effectively quench the intrinsic fluorescence of BCI through static quenching. The stoichiometric binding between the two was observed at a 1∶1 ratio, and electrostatic interaction was the main force. Conclusion: A novel potato I-type chymotrypsin inhibitor with superior stability was developed using site-directed mutagenesis technology. This work expands upon the structure-function relationship studies of potato I-type protease inhibitors, providing a theoretical basis for their further development and application.
The complex and precise interactions between protein molecules are fundamental to various biological processes such as gene synthesis, cell proliferation, signal transduction, and cellular metabolism. These interactions play a key role in maintaining normal function and life activities of organisms. Based on synthetic biology techniques, researchers can effectively regulate these biomolecular interactions through modifying amino acids at protein interfaces or redesigning protein structures, thereby promoting the development of biotechnological fields including targeted peptide design and protein drug development. Compared with traditional experimental-based research methods, bioinformatics has demonstrated significant advantages in the study of interacting proteins, such as lower cost and higher efficiency. This paper systematically reviews the latest progress in the application of bioinformatics to the analysis, modification, and de novo design of interacting proteins. Based on a comparative analysis and systematic summary of existing research methods and strategies, a complete research framework for interacting proteins has been constructed, aiming to provide a reference for the translational application of interacting proteins.
Methotrexate (MTX) is an antagonist of the essential nutrient folic acid, which is widely used in the treatment of tumors and immune-related diseases with remarkable clinical efficacy. However, the majority of patients treated with MTX tend to experience organ toxicity, which severely limits the clinical use of MTX. More and more studies have shown that single nucleotide polymorphisms (SNPs) of some MTX metabolism genes are closely related to the metabolism and toxicity of MTX. In recent years, genetic testing based on premedication can determine personalized treatment options, so that patients can receive more effective treatment. Therefore, it is more important to understand the mechanism of MTX drug metabolism and elucidate the relationship between metabolic pathway-related gene SNPs and their adverse reactions. This study systematically introduces the metabolic pathways of MTX, reviews the relationship between its toxic side effects and SNPs of key genes, and discusses the precision medication of MTX, in order to provide references and suggestions for the personalized use of MTX in clinical practice.
Hydrogel matrix stiffness, as an important biomechanical factor, plays a crucial role in cell growth, migration, and differentiation. Studies have shown that the stiffness of hydrogels can significantly regulate cell-matrix interactions, thereby affecting cell proliferation, differentiation, and functional expression. Therefore, precisely controlling hydrogel stiffness to optimize cellular responses has become a key issue in tissue engineering and regenerative medicine. Hydrogel stiffness is closely related to its molecular structure, crosslinking density, and external environmental factors. Changes in stiffness can influence multiple biological processes, such as cell morphology, adhesion, proliferation, differentiation, and gene expression, through mechanical signaling pathways. Additionally, the effects of hydrogel stiffness vary among different cell types, making hydrogel-based cell culture and tissue construction a promising approach for various applications. This review summarizes the stiffness thresholds of hydrogels under different crosslinking systems and reveals the heterogeneous mechanisms of cellular mechanical sensitivity under various conditions, providing a theoretical foundation for future applications in regenerative medicine, cell engineering, and disease modeling.
The increasing global incidence of inflammatory bowel disease (IBD) poses a significant treatment burden on patients. Although traditional drug therapies such as aminosalicylic acid preparations and glucocorticoids dominate the treatment of IBD, problems such as drug resistance and side effects are becoming increasingly prominent, which limits their clinical efficacy. In recent years, the potential role of macrophages in the treatment of IBD has gradually attracted widespread attention. With the in-depth study of the mechanism of macrophage polarization and the emergence of novel drug delivery systems, targeting macrophage polarization to restore immune balance has become a promising therapeutic strategy. This paper systematically analyzes the types of macrophage polarization and its key role in intestinal homeostasis, and deeply explores the signaling pathways of macrophage polarization regulation in the treatment of IBD, as well as various ways of regulating macrophage polarization, including the regulation of traditional Chinese medicine, the application of new composite materials, and the alleviation of inflammation by changing the direction of macrophage polarization through microbiota metabolites. These research results provide not only new targets for the treatment of IBD, but also an important theoretical basis for the optimization and innovation of IBD treatment strategies.
Oil bodies are organelles found in microorganisms and animal and plant cells. They have important lipid storage and energy metabolism functions, and their unique structures are widely used in the food, pharmaceutical, and cosmetic industries. The oil body expression system simplifies the purification process for exogenous proteins and improves expression levels by fusing target proteins with oil body proteins, while the oil body delivery system takes advantage of the natural structure of oil bodies to address the safety and bioavailability issues of traditional delivery systems. This paper provides a comprehensive review of these two systems and offers some suggestions for meeting the needs of modern industries.
Isoprenoids have biological activities such as antioxidant, anti-inflammatory, and anti-cancer functions, and have important application value in food, medicine, cosmetics, and more. Currently, isoprenoids are often obtained through methods such as plant extraction and chemical synthesis, which lead to problems such as low yields and high cost in production. Diatoms are abundant, grow quickly and are widely distributed. They contain rich bioactive compounds, which have great potential as a biosynthetic platform for isoprenoids. The biological functions, synthesis, influencing factors and application prospects of several common isoprenoids in found diatoms are reviewed. The goal is to further study the role value of isoprenoids from diatoms, expand their applications in production and life, develop diatoms as a biosynthesis platform of isoprenoids, and provide a reference for the future industrialization of marine diatoms.
Biomedicine represents a strategically emerging industry,driven by cutting-edge technology and characterized by significant, sustainable growth potential. China’s biopharmaceutical sector has experienced rapid development, establishing the country as the world’s second-largest pharmaceutical market. Nevertheless, when compared to developed nations, certain limitations persist, including insufficient original innovation capabilities, inadequate policy coordination and interaction, limited motivation to transform scientific and technological achievements, and low efficiency in this regard. To address these challenges, we should draw from the successful experiences of developed countries in research and development, enterprise management, and policy-making. Efforts should focus on three key areas: enhancing the supply of original innovation outcomes, improving the evaluation and application system for innovation value, and optimizing the policy framework for the transformation of scientific and technological achievements. By eliminating bottlenecks and obstacles in the transformation process,we can promote quality enhancement and efficiency improvement within the biopharmaceutical industry, thereby accelerating the formation of new high-quality productive forces.
Currently, the major countries are prioritizing biomanufacturing as a key area of competition, and China is also actively fostering the biomanufacturing industry as a new engine of economic growth. Bioreactors are considered the core equipment in the biomanufacturing field and are referred to as the “heart” of the biomedical and biotechnology industries. With the continuous advancement of biotechnology, significant achievements have been made in bioreactors. Based on a comprehensive analysis of all research papers and patent literature in the field of bioreactors, this paper systematically reviews the current global technological development trends in the bioreactor domain. The study found that both scientific discoveries and technological inventions in the field of bioreactors are showing an upward trend. Scientific discoveries primarily focus on areas such as membrane bioreactors, which have become research hotspots, while technological inventions concentrate on solving issues like wastewater treatment. The Chinese Academy of Sciences ranks first in terms of scientific output, and domestic universities have significant influence. In terms of patents, domestic universities are the primary holders, yet the level of technology commercialization remains insufficient. Scientific research collaboration is characterized by regional cooperation, while patent collaboration is more scattered. Currently, both China and the United States are leaders in scientific research output in this field, and China is gradually gaining a late-mover advantage. Although China possesses a large number of technological patents, its number of Patent Cooperation Treaty(PCT) and tripartite patents is significantly lower than that of the United States, indicating the need to accelerate the development of core technological strengths.
