Biochemical Engineering Journal最新文献

{"title":"Expression and biological activity of recombinant human coagulation factor X in HEK293 cells","authors":"Sen Zou ,&nbsp;Jianing Feng ,&nbsp;Zhilan Zhang ,&nbsp;Shuai Fan ,&nbsp;Zhifei Zhang ,&nbsp;Zhaoyong Yang","doi":"10.1016/j.bej.2025.109939","DOIUrl":"10.1016/j.bej.2025.109939","url":null,"abstract":"<div><div>Recombinant human coagulation factor X (rhFX) plays a crucial role in hemostasis and holds potential therapeutic applications for bleeding disorders. This study aimed to establish an efficient expression and purification system for rhFX using HEK293 cells, enhancing its therapeutic potential. Employing molecular cloning and transient transfection, the FX gene was successfully integrated into the pcDNA3.1 vector and transfect to HEK293 cells, yielding high expression levels of rhFX, as confirmed by Western blot analysis and prothrombin time assays. The optimal conditions for protein expression were identified, with peak rhFX concentrations reaching ∼0.005 mg/L and demonstrating procoagulant activity of 5.79 %-6.30 %. Stable HEK293 cell lines were generated, showing consistent expression of rhFX and significantly increased procoagulant activity (61.1 %-78.3 %). Further adaptation to suspension culture improved rhFX yield, achieving concentrations of 13.16 mg/L and enhanced procoagulant activity (658.4 %). Safety assessments confirmed the absence of microbial contamination and genetic fidelity of cell lines. Additionally, the study established a robust screening model for FXa inhibitors, achieving an IC50 value for Edoxaban of 3.56 nM, which aligns with existing literature. Our findings highlight the successful production and biological validation of rhFX, emphasizing its potential for therapeutic applications in coagulation disorders.</div></div>","PeriodicalId":8766,"journal":{"name":"Biochemical Engineering Journal","volume":"225 ","pages":"Article 109939"},"PeriodicalIF":3.7,"publicationDate":"2025-09-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145118695","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Co-production of lactic acid and 2,3-butanediol by genetically engineered Enterobacter cloacae","authors":"Xinying Sun ,&nbsp;Wei Xiao ,&nbsp;Mengying Wu ,&nbsp;Fuqiang Liu ,&nbsp;Jiaxin Li ,&nbsp;Lei Liu ,&nbsp;Jing Wu ,&nbsp;Keke Cheng ,&nbsp;Jianan Zhang","doi":"10.1016/j.bej.2025.109938","DOIUrl":"10.1016/j.bej.2025.109938","url":null,"abstract":"<div><div>Microbial fermentation technology has advanced rapidly in the field of green chemical production, showcasing broad application prospects. However, current techniques still face challenges such as low carbon utilization efficiency and excessive accumulation of by-products. Additionally, during the fermentation process, carbon sources are released in the form of CO₂, leading to significant carbon loss. In this study, <em>Enterobacter cloacae</em> CICC 10011 was metabolically engineered to achieve the co-production of 2,3-butanediol (2,3-BDO) and lactic acid, while reducing carbon dioxide emissions and improving carbon utilization. By knocking out key genes (<em>pflB</em> and <em>iclR</em>), the pathways for the formation of by-products such as formate, acetate, and ethanol were attenuated, thereby optimizing carbon flux toward the target products. The results showed that under optimal fermentation conditions (aeration rate of 0.4 vvm, pH 6.5, temperature 35 °C), the double-gene deletion strain ECΔpflBΔiclR achieved lactic acid and 2,3-BDO (mixture of stereoisomers) concentrations of 51.03 g/L and 10.44 g/L, respectively, with a total target product yield of 1.76 mol/mol. Compared to the wild-type strain, the production of by-products succinate, acetate, and ethanol was reduced by 70.04 %, 47.10 %, and 89.76 %, respectively. Additionally, CO₂ emissions were reduced by 68.92 %, and carbon conversion efficiency improved by 116.16 %. This strategy provides a novel approach for carbon emission reduction and the co-production of high-value chemicals in fermentation processes.</div></div>","PeriodicalId":8766,"journal":{"name":"Biochemical Engineering Journal","volume":"225 ","pages":"Article 109938"},"PeriodicalIF":3.7,"publicationDate":"2025-09-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145118694","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Nitrogen removal performance and microbial community dynamics in simultaneous anammox-denitrification filter column under tetracycline stress","authors":"Ting Sun ,&nbsp;Yongbao Pan ,&nbsp;Xiyan Du ,&nbsp;Chunyan Ren ,&nbsp;Qian Huang ,&nbsp;Huihui Zhang ,&nbsp;Chongyang Wang ,&nbsp;Yong-tao Lv","doi":"10.1016/j.bej.2025.109937","DOIUrl":"10.1016/j.bej.2025.109937","url":null,"abstract":"<div><div>Antibiotic accumulation causes serious environmental and health risks. However, its impact on the performance of anaerobic ammonium oxidation (anammox)-based nitrogen removal system remains poorly understood. In this study, a simultaneous anammox and denitrification (SAD) filter column reactor was constructed and the long-term effects of tetracycline (TC) on its total nitrogen (TN) removal efficacy and microbial community dynamics were investigated. Results demonstrated that the SAD system achieved a TN removal efficiency of 86.2 % with a low nitrate concentration in effluent (3.73 mg·L⁻¹). At 0.05–0.2 mg·L⁻¹ TC, the TN removal efficiency of system increased to 95.6 %. However, 0.5 mg·L⁻¹ TC induced short-term inhibition of the activity of anammox bacteria (AnAOB) and the TN removal efficiency of system decreased to 87.8 %. However, the TN removal efficiency of system recovered to 95.9 % after 21 days. Further analysis revealed that TC stress increased the particle size of granular sludge to 536.5 μm and elevated the proportion of tightly bound extracellular polymeric substances (TB-EPS) to 63.7 %. Microbial community analysis revealed spatial heterogeneity, with the bottom layer (0–15 cm) being enriched with <em>Candidatus</em> Kuenenia (12.7 %) and the middle layer (15–35 cm) being dominated by the denitrifying bacteria <em>Denitratisoma</em> (37.4 %). This study provides a theoretical basis and process optimization strategy for efficient treatment of antibiotic-rich wastewater via SAD process.</div></div>","PeriodicalId":8766,"journal":{"name":"Biochemical Engineering Journal","volume":"225 ","pages":"Article 109937"},"PeriodicalIF":3.7,"publicationDate":"2025-09-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145096032","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Effect of dynamic culture conditions on the production of an erythroid progenitor cell line within fluidised bed bioreactors","authors":"David Phillips ,&nbsp;Marianne J. Ellis ,&nbsp;Jan Frayne ,&nbsp;Sandhya Moise","doi":"10.1016/j.bej.2025.109936","DOIUrl":"10.1016/j.bej.2025.109936","url":null,"abstract":"<div><div>Red blood cells (RBCs) are an essential therapeutic resource; however, insufficient supply and risks of blood-borne infections drive the need for alternative sources. The Bristol Erythroid Lineage – Adult (BEL-A) erythroid progenitor cell line provides a new source for the indefinite production of cultured RBCs, but efficient and scalable bioprocessing strategies are yet to be established. Fluidised bed bioreactors (FBBs), with their low-shear environments and high mass transfer capabilities, can support high-density cultures, particularly at large scales, offering a promising manufacturing platform for BEL-A expansion. However, their optimal operating conditions and effects on BEL-A cell have yet to be defined. Using a design of experiments statistical approach, we systematically investigated how dynamic culture conditions impact BEL-A proliferation within FBBs. A lower media perfusion velocity, reduced initial cell seeding number, and a higher cell density (cells/mL) enhanced cellular proliferation. We demonstrate that FBB culture achieved productivity comparable to static culture whilst offering scalability and reduced manual handling. Importantly, no spontaneous differentiation of BEL-A cells was observed, confirming the system’s suitability for maintaining progenitor cell characteristics. This study is the first to demonstrate the feasibility of FBBs for mammalian single-cell suspension culture, using BEL-A as a model system. Furthermore, our work represents a critical step towards the clinical-scale manufacture of BEL-A cells and unlocking their therapeutic potential as a source of cultured RBCs.</div></div>","PeriodicalId":8766,"journal":{"name":"Biochemical Engineering Journal","volume":"225 ","pages":"Article 109936"},"PeriodicalIF":3.7,"publicationDate":"2025-09-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145096037","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Remediation mechanism of Sporosarcina pasteurii XL-1 for removal of Cr(VI) and Cr(III) by induced carbonate precipitation","authors":"Yanli An,&nbsp;Xilin Li,&nbsp;Ling Liu,&nbsp;Jijia Liu,&nbsp;Ying Yu,&nbsp;Jiahong Zhang,&nbsp;Zhongkai Tong","doi":"10.1016/j.bej.2025.109933","DOIUrl":"10.1016/j.bej.2025.109933","url":null,"abstract":"<div><div>In this study, a strain capable of inducing carbonate precipitation was screened and obtained from the polluted soil near the chromium slag dump; A chromium ion tolerance domestication test was conducted to determine the bacterial identification (optical density (OD₆₀₀), pH, electrical conductivity (EC), urease activity(UA), NH₄⁺ ); Reduction and mineralisation kinetics, combined with scanning electron microscope (SEM), energy dispersive spectrometer (EDS), fourier transform infrared spectroscopy (FTIR), and X-Ray diffraction (XRD) were used to analyse the removal mechanism of Cr(III) and Cr(VI). The strain was identified as <em>Sporosarcina pasteurii</em> XL-1; with maximum tolerance level of 1200 mg/L for Cr(III) and 500 mg/L for Cr(VI). Reduction kinetics indicated that Cr(VI) reduction was primarily by extracellular enzymes; Mineralisation kinetics indicate that the bacterial solution could regulate the crystal morphology, leading to gradual aggregation and bonding that resulted in a denser mineral structure favorable for the encapsulation and removal of heavy metals such as chromium; Microscopic analysis showed Cr(III) removal mechanism included bio-induced carbonate precipitation and abiotic precipitation of Cr(OH)₃; Cr(VI) removal mechanism was mainly bioreduction, supplemented by bio-induced carbonate precipitation and abiotic Cr(OH)₃ precipitation. Finally, a process model was proposed to describe the mineralisation and immobilisation of Cr(VI) and Cr(III) via MICP, offering theoretical guidance for the subsequent remediation of chromium-contaminated water.</div></div>","PeriodicalId":8766,"journal":{"name":"Biochemical Engineering Journal","volume":"225 ","pages":"Article 109933"},"PeriodicalIF":3.7,"publicationDate":"2025-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145096030","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"De Novo biosynthesis of scopoletin from glucose by engineered Saccharomyces cerevisiae","authors":"Junjie Wang,&nbsp;Pengming Yang,&nbsp;Shuai Tu,&nbsp;Weihong Zhong","doi":"10.1016/j.bej.2025.109934","DOIUrl":"10.1016/j.bej.2025.109934","url":null,"abstract":"<div><div>Scopoletin, a plant-derived coumarin, exhibits diverse pharmacological activities. Nowadays, scopoletin is mainly produced by plant extraction. In this study, a <em>Saccharomyces cerevisiae</em> strain capable of <em>de novo</em> scopoletin biosynthesis was constructed by successively integrating ten heterologous genes into a <em>p</em>-coumaric acid producing strain. The following strategies were employed to improve the yield of scopoletin: (1) A high-yield caffeic acid platform strain (301.98 mg/L) was established by increasing the <em>HpaBC</em> copy number, integration of <em>EcaroL</em> and <em>RgTAL</em>, feedback-resistant mutant expression cassette (<em>ARO3</em>^{<em>D154N</em>}/<em>ARO4</em>^{<em>G141S</em>}/<em>ARO7</em>^{<em>K229L</em>}), and knockout competing pathway genes (<em>ARO10</em>/<em>PDC5</em>); (2) Two distinct pathways from caffeic acid to scopoletin were introduced and selected; (3) Integration of heterologous genes <em>AtHCT</em> and <em>AtC3’H</em>, to construct the pathway from <em>p</em>-coumaroyl-CoA to caffeoyl-CoA; (4) Attenuating <em>PYK1</em> by introducing the mutant gene of <em>PYK1</em>^{<em>D146N</em>} to increase PEP supply; (5) Overexpression of cofactor regeneration genes (<em>ADO1</em>, <em>SAH1</em>, and <em>ZWF1</em>) to strengthen the supply of SAM and NADPH; (6) Adjustment of the fermentation medium. The engineered strain produced scopoletin of 55.32 mg/L in shake flasks. To our knowledge, this is the first report of <em>de novo</em> scopoletin biosynthesis in <em>S. cerevisiae</em>. These results establish new insights for microbial production of scopoletin and coumarin derivatives.</div></div>","PeriodicalId":8766,"journal":{"name":"Biochemical Engineering Journal","volume":"225 ","pages":"Article 109934"},"PeriodicalIF":3.7,"publicationDate":"2025-09-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145096031","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A novel Fe(II) dosing strategy for a self-sustaining microalgal-bacterial membrane photobioreactor: Performance improvement and fouling mitigation","authors":"Zhaozhao Wang ,&nbsp;Yaxin Wang ,&nbsp;Lina Yan ,&nbsp;Dameng Lian ,&nbsp;Baoqiang Liao","doi":"10.1016/j.bej.2025.109932","DOIUrl":"10.1016/j.bej.2025.109932","url":null,"abstract":"<div><div>In microalgal-bacterial (MB) consortia, the production and exchange of O₂ and CO₂ between microalgae and bacteria have the potential of completely eliminating external aeration and carbonation and thus creating a self-sustainable MB ecosystem for simultaneous chemical oxygen demand (COD), N, and P removal in a single step. The self-sustaining microalgal-bacterial membrane photobioreactor (SSMB-MPBR) has attracted increasing attention due to its energy conservation and environmental friendliness. Nevertheless, membrane fouling caused by microalgae proliferation significantly reduces operational sustainability. This study examines the impact of Fe(II) dosing on pollutant removal and fouling control of an SSMB-MPBR process in treating municipal wastewater. The results show that on average, 92.77 % of chemical oxygen demand (COD), 76.34 % of total nitrogen (TN), and 89.86 % of total phosphorus (TP) were removed with zero external aeration, representing improvements of 2.61 %, 5.80 %, 8.10 %, respectively, due to increased biomass productivity (75 mg/(L·d)) after Fe(II) dosing. Additionally, the membrane fouling rate (F_r) was reduced by 28.60 % because of lower extracellular polymeric substances (EPS) levels and larger microbial aggregates, which formed a loose biocake. Illumina sequencing results showed that Fe(II) dosing enriched functional microalgae and optimized the interaction between microalgae and bacteria. These findings demonstrate that Fe(II) dosing is a practical strategy to improve treatment performance and mitigate membrane fouling of the SSMB-MPBR process.</div></div>","PeriodicalId":8766,"journal":{"name":"Biochemical Engineering Journal","volume":"225 ","pages":"Article 109932"},"PeriodicalIF":3.7,"publicationDate":"2025-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145044739","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Alkali pretreatment as a game-changer in enhancing methane production from cattle manure and reshaping microbial landscape: A comparative analysis of NaOH, KOH, and Ca(OH)2","authors":"Wenjun Yan ,&nbsp;Zhixuan Yin ,&nbsp;Lingying Kong ,&nbsp;Qian Zhang ,&nbsp;Changqing Liu","doi":"10.1016/j.bej.2025.109931","DOIUrl":"10.1016/j.bej.2025.109931","url":null,"abstract":"<div><div>The presence of refractory organic substances in cattle manure significantly impedes its biogas recovery potential through anaerobic digestion. To address this challenge, this study explored the application of three different types of alkaline agents—NaOH, KOH, and Ca(OH)₂—for pretreatments to optimize methane production from cattle manure. The results demonstrated that pretreatment with various alkaline agents could notably accelerate the conversion of insoluble organics in cattle manure into more digestible soluble forms, with varying degrees of effectiveness. NaOH/KOH pretreatments of cattle manure elevated the soluble chemical oxygen demand (SCOD) by 7.6 times, reaching approximately 38,000 mg/L. This enhancement led to a 50 % increase in the methane yield during subsequent anaerobic digestion, achieving a rate of 150 mL/g VS·d. Despite the low solubility of Ca(OH)₂ and the formation of Ca-precipitates hindering hydrolysis during pretreatment, the methane yield still increased by 31 %. High-throughput sequencing further revealed that alkaline pretreatments reshaped the microbial communities. Specifically, by enhancing hydrolysis and increasing the availability of H₂/CO₂, NaOH/KOH pretreatment shifted the methanogenesis pathway from acetotrophic to polytrophic by enriching hydrogenotrophic <em>Methanobacterium</em> and polytrophic <em>Methanosarcina</em>. However, Ca(OH)₂ pretreatment maintained the dominance of acetoclastic methanogenesis by preserving the inherent characteristics of the cattle manure.</div></div>","PeriodicalId":8766,"journal":{"name":"Biochemical Engineering Journal","volume":"225 ","pages":"Article 109931"},"PeriodicalIF":3.7,"publicationDate":"2025-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145045609","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"An elastin-like polymer targeting vascular endothelial growth factor receptor-1 reduces survival in serum-starved endothelial cells","authors":"M. Lisa Phipps ,&nbsp;Antonietta M. Lillo ,&nbsp;Demosthenes P. Morales ,&nbsp;Miriam Hernandez-Romero ,&nbsp;Leyma P. De Haro ,&nbsp;Devin Close ,&nbsp;Wynter A. Paiva ,&nbsp;Emily K. Funsten ,&nbsp;Andrew R.M. Bradbury ,&nbsp;Jennifer S. Martinez ,&nbsp;Eva Rose M. Balog","doi":"10.1016/j.bej.2025.109930","DOIUrl":"10.1016/j.bej.2025.109930","url":null,"abstract":"<div><div>Peptides often exhibit biological activity that depends on the context in which they are displayed and delivered. Understanding and controlling these contextual effects on peptide function is critical for designing targeted and responsive peptide-based biomaterials and therapeutics. Genetically engineered protein polymers such as elastin-like polypeptides (ELPs) can incorporate bioactive peptide motifs and are attractive candidates for biomaterials used in tissue engineering and targeted drug delivery. They also present an opportunity for investigating and modulating cell signaling pathways by presenting a peptide ligand in various defined chemical and physical environments. Vascular endothelial growth factor receptor-1 (VEGFR1) signaling plays important and complex roles in cell survival and angiogenesis, but polymeric materials that interact with this signaling axis are scarce. In this study, a novel genetically engineered elastin-like polymer that targets VEGFR1 is characterized. This polymer, termed R1B-ELP, binds to human endothelial cells in a manner dependent on its VEGFR1-targeting motif and, based on cell proliferation and cytotoxicity assays, demonstrates activity consistent with disrupting pro-survival signaling necessary for endothelial cell function under conditions of environmental stress. Notably, these findings indicate that ELP fusion alters the functional behavior of the targeting peptide. Modulators of VEGFR1 signaling have potential applications in basic studies of angiogenesis as well as in therapeutic applications targeting vascular or inflammatory diseases.</div></div>","PeriodicalId":8766,"journal":{"name":"Biochemical Engineering Journal","volume":"225 ","pages":"Article 109930"},"PeriodicalIF":3.7,"publicationDate":"2025-09-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145045597","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Methylcellulose in culture medium regulates size, cell viability, cytokine production, and exosome secretion in adipose-derived stem cell spheroids cultured using SphereRing® system","authors":"Takuya Sakamoto ,&nbsp;Hiroto Koma ,&nbsp;Ayane Kuwano ,&nbsp;Tetsuhiro Horie ,&nbsp;Atsushi Fuku ,&nbsp;Hironori Kitajima ,&nbsp;Yuka Nakamura ,&nbsp;Ikuhiro Tanida ,&nbsp;Yujiro Nakade ,&nbsp;Yoshiyuki Tachi ,&nbsp;Ikki Horiguchi ,&nbsp;Naoki Yamamoto ,&nbsp;Sohsuke Yamada ,&nbsp;Xin Guo ,&nbsp;Qian Yang ,&nbsp;Yasuhito Ishigaki ,&nbsp;Toru Ichiseki ,&nbsp;Ayumi Kaneuji ,&nbsp;Satoshi Osawa ,&nbsp;Norio Kawahara","doi":"10.1016/j.bej.2025.109929","DOIUrl":"10.1016/j.bej.2025.109929","url":null,"abstract":"<div><div>Knee osteoarthritis (OA) is a degenerative disease that impairs joint function. Adipose-derived stem cells (ADSCs) are promising for therapy due to their regenerative potential, particularly when cultured as spheroids to enhance paracrine activity. However, conventional plate-based spheroid culture faces challenges such as scalability limits, retrieval difficulties, excessive enlargement, and necrotic core formation, all of which may compromise therapeutic efficacy. An optimized, scalable method is therefore required for clinical application. This study aimed to establish a spheroid culture protocol using the SphereRing® device with methylcellulose (MC) to regulate spheroid size, improve viability, and enhance secretion of therapeutic factors including exosomes and interleukin-10 (IL-10). ADSCs were cultured in SphereRing® with 0.5–1 % MC for 3 days, and spheroid size, morphology, viability, and secretory profiles were assessed. Spheroids cultured with 0.75 % MC exhibited the most uniform size distribution, higher circularity, reduced necrotic core formation, and significantly greater viability compared to untreated controls. Moreover, IL-10 and exosome secretion were markedly increased in the 0.75 % MC group. Patient-derived ADSCs showed comparable improvements. MC-treated spheroids also maintained superior viability after exposure to synovial fluid, simulating intra-articular conditions. These findings suggest that incorporating MC into the SphereRing® system enables scalable and uniform spheroid production, establishing a clinically relevant platform for ADSC-based therapy in knee OA with potential to improve treatment consistency and outcomes.</div></div>","PeriodicalId":8766,"journal":{"name":"Biochemical Engineering Journal","volume":"225 ","pages":"Article 109929"},"PeriodicalIF":3.7,"publicationDate":"2025-09-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145045610","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}