Hypertrophic scars have long posed a significant challenge in wound healing, primarily because of an imbalance between Type I (COL-I) and Type III (COL-III) collagen caused by excessive fibroblast activation. Existing treatments cannot directly regulate collagen formation to intervene in scar hyperplasia. This study is the first to elucidate the key mechanism through which human umbilical cord-derived mesenchymal stem cell (hUC-MSCs) exosomes expressing miR-185-5p achieve scarless healing through remodeling of the collagen-type ratio. In this method, a hypoxic three-dimensional (3D-HO) suspension culture system that more closely mimics the in vivo microenvironment was established, and functional exosomes with enhanced physiological activity and higher yields were harvested. In vivo experiments demonstrated that 3D-HO exosomes significantly improved the COL-III/COL-I regeneration ratio during mouse wound healing. Mechanistic studies revealed that the RhoA/YAP signaling axis plays a key regulatory role in the collagen regeneration ratio. Further molecular analysis revealed enrichment of miR-185-5p in 3D-HO exosomes, which directly target RhoA to regulate fibroblasts. Both in vivo and in vitro functional interventions confirmed miR-185-5p in maintaining collagen type balance. In summary, this study demonstrates that microenvironment-enhanced hUC-MSCs exosomes remodel the composition of different types of collagen via the miR-185-5p-RhoA/YAP signaling axis, thereby driving scarless healing.

Chinese hamster ovary(CHO)cells are the most widely used hosts for biopharmaceutical production,yet their low recombinant antibody expression efficiency remains one of key bottlenecks in industrial applications.To enhance anti-EGFRv III antibody expression in CHO cells,an intensified fed-batch process was successfully developed for Cell A and Cell B cell strains based on cell cycle regulation.First,flow cytometry was used to establish the relationship between cell cycle distribution and specific antibody production rate(Qp).The results showed that the proportions of cells in the G0/G1 phase increased significantly over time,reaching 50%and 64%on day 12 for Cell A and Cell B,respectively.Meanwhile,the single-cell Qp increased by 1.5 folds(Cell A)and 1.0 fold(Cell B)compared with initial levels.Higher proportions of cells in the G0 and G1 phases could significantly boost Qp,however,impair cell growth and accumulation,leading to a reduction in total antibody titer.To address this,we implemented feeding during the N-1 seed stage and extended the culture time.At harvest,the densities of Cell A and Cell B reached 26.3×106 cells/mL and 17.7×106 cells/mL,with the proportions of cells in G0/G1 phase increasing to 45%and 38%,respectively.After inoculation of high-density seed cultures into 5 L bioreactors and continuing culture for 14 days,the integrated viable cell density(IVCD)of both Cell A and Cell B exceeded that of the conventional process.In addition,the treatment increased the Qp by 11.0%and 9.1%and achieved final antibody titers of 9.5 g/L and 5.0 g/L for Cell A and Cell B,respectively,with no significant impact on critical quality attributes of recombinant antibody.In summary,the intensified fed-batch process established in this study for CHO cells,based on cell cycle regulation,simultaneously enhances cell accumulation and specific productivity,ultimately significantly improving the recombinant antibody expression levels in CHO cells.Therefore,this intensified fed-batch process can be adopted for recombinant antibody production by CHO cells cultivated at an industrial scale.

To address the challenges of high-density animal cell culture, this study developed a high-density culture system comprising a single-use bioreactor (SUB) with a nominal volume of 500 mL and a pulsed tangential flow filtration (ITF) unit. The reactor can be configured with two layers of 35 mm diameter impellers—either double Elephant Ear (EE-EE) or Elephant Ear combined with Ribbon (EE-RB). The flow field characteristics were rigorously characterized through CFD simulations (60–240 rpm; 90–480 mL) validated by experimental data. Engineering analysis revealed the system’s robust culture environment: k_La values ranging from 2 to 15 h−1 (60–180 rpm; 200–400 mL; 30–150 mL/min aeration), P/V values from 0.1 to 10 W/m3, and mixing times between 1 and 10 s. Crucially, the system maintains a mild shear environment with an average shear strain rate (SSR) below 25 s−1, fully within the physiological tolerance range for mammalian cells. This low-shear, high-mass-transfer design was validated through perfusion cultures of CHO and HEK293 cells. The system achieved exceptionally high cell densities of 8.32 × 107 cells/mL and 1.17 × 108 cells/mL, respectively, with cell viability consistently exceeding 90

Promoting rapid wound healing is essential for preventing hypertrophic scars. Exosome-based therapies represent a promising approach for improving healing outcomes, but they encounter significant challenges: 1) the isolation of exosomes requires complex, time-consuming, and costly procedures; 2) the stability of exosomes is limited due to the pH-and temperature-sensitive nature of their lipid bilayer. Hydrogels, known for their hydrophilic and porous architectures, offer an attractive solution by enabling the isolation of exosomes from macromolecular impurities while simultaneously protecting them from degradation. In this study, we designed a bilayer polysaccharide hydrogel where pullulan (PL) and calcium ions (Ca) form the inner core, while sodium alginate (SA) creates an effective isolating gel in the exosome-containing precursor solution. By integrating freeze-drying with ultrasonic-assisted dissolution, the approach effectively achieves both the isolation and controlled release of exosomes. The results show that the PL + Ca + SA hydrogel exhibits superior gelation stability compared to other formulations. During isolation, the PL + Ca + SA hydrogel efficiently enriched exosomes while maintaining their characteristic "teacup-shaped" morphology and protein markers. The isolation yield (87.2%) was comparable to that obtained by ultracentrifugation, with no significant differences in particle number or impurities. Furthermore, ultrasonic-assisted dissolution enabled sustained exosome release for at least 7 days, as predicted by the Higuchi diffusion model. Functionally, the EXO@PL + Ca + SA hydrogel enhanced fibroblast proliferation and viability for up to one week, changing the ROS content (decreasing 92%) and the COL-I/COL-III ratio to 0.58 of mouse wound model. Furthermore, transcriptomic analysis revealed significant alterations in related pathways involving key genes. Overall, this bilayer hydrogel offers an integrated platform for exosome isolation and controlled delivery, providing preliminary experimental evidence to support further translational research.

In this study, Se-doped CeO2@Fe3O4 nanoparticles (NPs) were synthesized and applied to a Carthamus tinctorius (safflower) cell suspension culture using liquid medium (B5). The application of these NPs at various levels (0, 5, 10, 15 and 20 mg L-1) was studied for its effects on cell growth, physio-biochemical traits and antioxidative activities. The addition of NPs to the culture media significantly improved the cell biomass, antioxidant potential and phenolic contents. The addition of NPs at the rate of 15 mg L-1 (T3 treatment group) significantly improved the dry biomass of cells (128.72%), total chlorophyll contents (76.02%), and reduced levels of hydrogen peroxide (5.15%) and reactive oxygen (26.51%) compared to the control group (0 mg L-1). Furthermore, this study identified 29 differentially expressed genes (DEGs) in the jasmonate signalling pathway. Notably, only the two DEGs from the MYC2 family showed mixed expression at different time points (6 h, 24 h, 48 h, and 72 h) following treatment with Se-doped CeO2@Fe3O4 NPs. In conclusion, these findings demonstrate that this approach is effective, adaptable, biocompatible, and cost-efficient, offering a promising strategy for enhancing the production of antioxidant and bioactive metabolites in industrial-scale safflower cultivation.

Organoid-based therapy, as an approach in regenerative medicine, offers new options for previously untreatable diseases. However, the limitations of organoid transplantation, such as immunogenicity, tumorigenic potential, and ethical issues, restrict its clinical translation. For the first time, a novel three-dimension matrigel-free suspension culture system is developed to generate liver ductal organoid-derived extracellular vesicles (3D OEVs) for primary sclerosing cholangitis (PSC) treatment. The results show that the developed suspension culture provides a more favorable mechanical microenvironment, which significantly enhances the functional maturation of liver ductal organoids and their extracellular vesicles. This improvement is facilitated by a multidimensional regulatory network that encompasses the PI3K-AKT and RAP1 pathways. Moreover, 3D OEVs significantly attenuate hepatic inflammation and fibrosis because they can remodel the immune microenvironment, polarizing more macrophages into the anti-inflammatory M2 phenotypic macrophages. Furthermore, miR-1299 enriched in 3D OEVs is identified as the core effector molecule for macrophage reprogramming, which was confirmed by both in vivo and in vitro experiments to effectively repair biliary damage through suppressing the EGR1/FOS/RAS signaling axis. As a result, a scalable platform for the production of therapeutic OEVs was successfully developed in a 50 mL bioreactor utilizing a matrigel-free methodology, thus offering a bioactive material for liver regeneration.

Intratumoral redox homeostasis is often implicated in the development of multidrug resistance (MDR), which compromises the oxidative cytotoxicity of anthracycline chemotherapeutics such as doxorubicin (DOX). To overcome the redox homeostasis-driven MDR, we analyzed the transcriptomes of 216 breast cancer patients, revealing a noteworthy association between the TRPA1 (a regulator of redox homeostasis) overexpression and drug resistance. Moreover, a smart magnetic nanozyme, HADAF (HA@Anti-TRPA1@DOX@Au@Fe3O4), was rationally designed to target TRPA1 and reverse MDR. HADAF nanozyme integrates Au@Fe3O4 nanosheets with DOX, antisense oligonucleotides of TRPA1 and hyaluronic acid (HA). Notably, the dynamic regulation of HADAF is triggered spatiotemporally by a controllable low-frequency vibrational magnetic field (VMF) and near-infrared (NIR) laser irradiation. In vitro and in vivo experiments showed that HADAF effectively inhibited the TRPA1/PI3K/mTOR/MCL-1 signaling pathway, promoting apoptosis and ferroptosis, and achieving a 90% inhibition rate in MCF-7/ADR cells. As a result of this engineered therapeutic approach, the combination therapy reduced the IC50 of DOX by 87.7-fold compared to free DOX. In summary, this multimodal nanozyme provides a novel strategy to target redox homeostasis and overcome MDR in cancer therapy.

Scalable biomanufacturing of exosomes is constrained by poorly defined hydrodynamic conditions that decouple process control from mesenchymal stem cell viability, metabolism, and exosome quality. In this study, a computational fluid dynamics (CFD)–guided shear-window engineering framework was established to enable robust, serum-free exosome manufacturing in a wave bioreactor. A spatially homogeneous low-shear operating window (0.08–0.19 Pa, kLa 4.9–6.5 h−1) was identified and implemented in three-dimensional serum-free culture of human umbilical cord mesenchymal stem cells. Operating within this window resulted in an approximately 15-fold increase in exosome yield while maintaining exosome size distribution and canonical marker expression. This enhancement was accompanied by a metabolically stabilized cellular state, characterized by increased ATP production, improved NAD+/NADH balance, reduced lactate accumulation, and Ca2+-associated mechanotransduction consistent with Piezo1-related signaling. Exosomes produced under these conditions exhibited preferential renal accumulation within 12–24 h after systemic administration and demonstrated robust antifibrotic efficacy across cellular, organoid, and diabetic kidney disease models. Collectively, quantitative shear-window engineering is established as a process-oriented paradigm for the scalable biomanufacturing of exosomes.

The immunosuppressive tumor microenvironment (TME) plays a critical role in the exacerbation of chemotherapy-induced multidrug resistance (MDR). To overcome MDR, a natural nanovesicle-based biomimetic nanosystem was created using low-frequency vibrational magnetic fields (VMF) and an 808 nm laser to help overcome drug resistance in tumors. This advanced platform integrates M1 macrophage-derived exosomes (M1-EXO) with magnetic nanoparticles, Ce6 labeled antisense oligonucleotides of heat shock protein 70 (HSP-70) and doxorubicin. This approach enables a triple-modality synergistic therapy that integrates immunotherapy, gene therapy, and enhanced photothermal therapy (PTT). Specifically, M1-EXO effectively reprogram tumor-associated M2 macrophages (TAMs) toward the antitumor M1 phenotype, while Ce6-mediated photodynamic therapy (PDT) amplifies reactive oxygen species (ROS)-dependent M1 repolarization. Further, the nanovesicles target HSP70 to suppress heat shock protein expression, thereby overcoming the thermal resistance of tumor cells and enhancing PTT. Concurrently, nanovesicles' favorable magnetic responsiveness enables direct destruction of cancer cells under VMF exposure, which greatly contributed to immunogenic cell death (ICD) activation. Consequently, this synergistic strategy initiates antitumor immunity and activates cytotoxic T lymphocytes. Both in vitro and in vivo studies demonstrate that this biomimetic nanovesicle reduces the half-maximal inhibitory concentration (IC50) of doxorubicin-resistant breast cancer cells by 96-fold. In conclusion, this nanoplatform successfully tackles drug resistance by actively targeting pathways that activate the immune system, and promotes exosomes from human peripheral blood mononuclear cells for transplantation immunotherapy in the future.

Mesenchymal stem cell-derived exosomes (MSCs-EXOs) have demonstrated significant therapeutic potential in cholestatic liver disease. However, clinical translation has been hindered by challenges in scalable production and the incomplete elucidation of the underlying mechanisms. Herein, we aimed to enhance both the production scalability and antifibrotic efficacy of MSCs-derived exosomes by utilizing exosomes produced through threedimensional bioreactor culture (3D-EXOs), and to elucidate their molecular mechanisms. In a murine model of cholestatic liver fibrosis, 3D-EXOs exhibited superior antifibrotic and anti-inflammatory efficacy compared with exosomes derived from two-dimensional culture (2D-EXOs). Hepatic fibrosis and pro-inflammatory cytokine expression were significantly attenuated following treatment with 3D-EXOs. Furthermore, 3D-EXOs induced a more pronounced suppression of M1 macrophage polarization markers in vitro. Integrated transcriptomic analysis with experimental validation revealed that miR-1291, highly enriched in 3D-EXOs, functions as a key mediator. miR-1291 was shown to inhibit M1 macrophage polarization by concurrently modulating the SP1/ MAPK3/MAPK and TRAF3/NF-kappa B signaling pathways, thereby attenuating inflammatory responses. In this study, we established an effective method to enhance exosome production and enrich miR-1291 content using a three-dimensional culture system, resulting in substantial improvement in antifibrotic therapy. The mechanism involves suppression of M1 macrophage polarization through a novel dual-pathway regulatory network. These findings provide critical insights for developing exosome-based therapeutics for hepatic diseases and underscore the need for further long-term investigations.

Chinese hamster ovary (CHO) cells are the primary platform for therapeutic antibody production. Although histone deacetylase inhibitors such as sodium butyrate (NaBu) can enhance recombinant expression, their growth-inhibitory and cytotoxic effects often limit volumetric productivity. Here, a structure-guided docking strategy was applied to prioritize NaBu-derived small-molecule additives (SMAs) for experimental screening in CHO antibody-producing cell lines. A lead combination (D1 +D4) increased volumetric antibody titers by approximately 2-3 fold while maintaining high cell viability (>95%) under the tested conditions. Cell-based analyses indicated reduced apoptotic markers and altered cell-cycle distributions in D1 +D4-treated cultures relative to NaBu. Metabolite profiling further revealed reduced by-product accumulation, including lactate and ammonia, together with a distinct intracellular energy and redox state compared with control cultures. Importantly, N-glycan profiles and charge variants of the produced antibodies remained comparable between control and D1 +D4-treated cultures. The productivity benefit of D1 +D4 was further maintained in bench-scale stirred-tank and wave bioreactor systems, supporting process relevance. Together, these results demonstrate that structure-guided NaBu analog screening can identify practical additives that enhance CHO antibody productivity with minimal impact on product quality.

Breast cancer poses a significant health threat to women, as traditional chemotherapy often fails due to multidrug resistance and tumor adaptability. Moreover, the anti-apoptotic nature of these tumors limits the effectiveness of conventional treatments. Current therapeutic approaches, including photodynamic therapy (PDT), face limitations in clinical application due to poor water solubility of photosensitizers and challenges in targeted delivery. This study aims to develop a multifunctional nanosystem that integrates PDT with gene therapy to enhance breast cancer treatment efficacy. Firstly, Fe3O4 hollow nanospheres were prepared to carry Ce6 drug to kill breast cancer cells via chemodynamic therapy and PDT, respectively. To overcome the drawback of poor water solubility of PDT, HA/G3139/Ce6@Fe3O4 nanosystem was synthesized, which combines the benefits of G3139 antisense oligonucleotides for gene therapy and hyaluronic acid for encapsulation and targeting CD44 receptor. The developed novel nanosystem exhibits excellent biocompatibility and pH responsiveness, significantly enhancing its cancer cell-killing capability under light exposure. When the therapeutic system enters cancer cells and decomposes in the acidic environment, it releases Ce6 and G3139. On one hand, the photosensitizer Ce6 generates cytotoxic ROS upon irradiation, killing cancer cells; on the other hand, G3139 binds to the anti-apoptotic gene BCL-2 in cancer cells, downregulating the protein and inhibiting tumor proliferation. The novel nanosystem demonstrates synergistic anti-cancer effects by combining PDT, gene therapy, and chemodynamic therapy, leading to enhanced apoptosis in breast cancer cells. In conclusion, this approach offers a promising strategy for more effective and targeted breast cancer treatment.

Elicitation is a potent strategy to enhance secondary metabolite yield, and nanoparticle-mediated elicitation offers a compelling alternative to traditional methods. Carthamus tinctorius (C. tinctorius) is rich in bioactive chlorogenic acids (CGAs) within its suspension cells. However, the mechanism by which doped nanoparticles (NPs) stimulate the CGAs biosynthesis in the cells remains unclear. Therefore, this study first elucidated the synthesis and characterization of selenium (Se)-doped CeO2@Fe3O4 NPs and then explored their regulatory effect and mechanism for the CGAs biosynthesis in C. tinctorius cells. Material characterization via TEM, XPS, XRD, and EDX analysis confirmed the successful synthesis of selenium-dopped CeO2@Fe3O4 nanocomposite. Fe3O4, CeO2, Fe3O4@CeO2, and selenium-dopped CeO2@Fe3O4 NP treatments all promoted the biosynthesis of CGAs in the cells. Notably, the group treated with 15 mg L-1 selenium-dopped CeO2@Fe3O4 NPs showed best effect (37.04 mg g-1 , 1.92 times that of control). Finally, metabolome and transcriptome profiling revealed the differential alterations in the content of 45 primary metabolites and differential expression of 83 CGA biosynthesis-related genes. This suggested that selenium-doped CeO2@Fe3O4 NPs up-regulated the expression of CGA biosynthesis- related genes, and consequently enhanced CGAs accumulation in the cells. This study provides the first evidence that selenium-doped CeO2@Fe3O4 acts as nano-elicitors for the biosynthesis of CGAs.

Introduction: Adipose stem cells (ADSC) have demonstrated therapeutic potential in ameliorating obesity and metabolic disorders, with their exosomes showing comparable therapeutic effects. However, the underlying molecular mechanism remains incompletely understood. Furthermore, the limited availability and inherent heterogeneity of primary ADSC present substantial challenges for consistent therapeutic outcomes. Objective: This study aimed to investigate the molecular mechanism underlying the unique biological effects mediated by microRNAs (miRNAs) in exosomes derived from immortalized adipose stem cells (iADSC). Methods and results: We first established stable iADSC and isolated their exosomes (iADSC-EXO), which exhibited both high yield and stability. In high fat diet (HFD)-fed obese mice, iADSC-EXO administration significantly attenuated obesity, reduced blood glucose and lipid levels, and alleviated hepatic steatosis. miRNA chips analysis revealed that miR-1246 is highly enriched in exosomes derived from iADSC and ADSC. Administration of miR-1246 to HFD-fed obese mice produced beneficial effects on obesity and metabolic disorders comparable to those with iADSC-EXO. Mechanistic studies revealed that miR-1246 exerts its beneficial effects through multiple pathways: First, reducing fat mass by downregulating of the expression of fat mass and obesity-associate protein (FTO) and subsequent inhibition of adipogenesis and lipogenesis. Second, enhancing white adipose tissue (WAT) beiging by suppressing Runt-related transcription factor 1 (RUNX1T1) and FTO expression. Third, promoting M2 macrophage polarization and suppressing WAT inflammation via inhibition of TNF receptor-associated factor 6 (TRAF6)-mediated inflammatory pathway. Conclusion: Our study elucidates a novel molecular mechanism through which ADSC regulates adipose tissue homeostasis and metabolism and presents a potential therapeutic approach for treating obesity and related metabolic disorders via iADSC-EXO or miR-1246-based interventions.

Halomonas elongata, a moderately halophilic γ-proteobacterium of industrial interest, serves as a microbial cell factory for ectoine-a high-value compatible solute extensively utilized in biopharmaceuticals and cosmetics. While its ectoine biosynthesis potential is well-documented, the systemic metabolic adaptations underlying osmoadaptation remain poorly characterized, limiting rational engineering strategies for optimized production. To address this gap, we employed chemostat cultivation coupled with multi-omics integration (physiological profiling, metabolomics, and metabolic flux analysis) to dissect salt-dependent metabolic network rewiring in the model strain DSM 2581T under moderate (6.0 % NaCl) and high salinity (13.0 % NaCl). Results demonstrated that, under moderate salt conditions, a specific growth rate (μ) of 0.20 h-1 significantly enhanced the ectoine-specific production rate (q p), intracellular ectoine content (p ectoine), and yield coefficient (Y p/s), concurrent with redirection of carbon flux toward the Entner-Doudoroff (ED) pathway and ectoine biosynthesis. Under high salt conditions, flux through both the ED pathway and ectoine biosynthesis was further upregulated, whereas fluxes through the pentose phosphate (PP) pathway, tricarboxylic acid (TCA) cycle, and CO2 generation were downregulated. Simultaneously, suppression of the flux from malate to pyruvate enhanced oxaloacetate synthesis, thereby increasing the supply of key precursors including glutamate, aspartate, and NADPH to fuel ectoine biosynthesis. Stepwise salt reduction experiments revealed bidirectional metabolic flexibility: elevated salinity prioritized carbon investment into ED-driven ectoine production, whereas hypo-osmotic conditions reactivate respiratory activity and the TCA cycle to fuel energy metabolism. These findings establish H. elongata as a paradigm of dynamic flux rewiring, where carbon economy is strategically reallocated between stress-protective solute biosynthesis and energy homeostasis. This study bridges the knowledge gap in understanding the physiological characteristics of H. elongata and provides a foundation for improving ectoine production and engineering strains through metabolic optimization.

Elevated succinate accumulation has been demonstrated to be associated with metabolic and inflammatory disorders. Our previous study revealed that adipose-derived stem cells (ADSC) from obese individuals exhibit high succinate, reduced biological activity, and mitochondrial dysfunction. However, the precise role of succinate in these processes remains unclear. Here, we investigated the effects of excess succinate on cellular biological activity, immunomodulatory capacity, and mitochondrial function of ADSC. We found that elevated succinate levels in ADSC decreased proliferation and differentiation potential, while promoting M1 macrophage polarization. Furthermore, succinate accumulation impaired mitochondrial biogenesis and metabolism, increasing in reactive oxygen species (ROS) production and inflammatory responses. Transcriptome sequencing analysis further confirmed that succinate upregulated inflammatory pathways, suppressed mitochondrial biogenesis and metabolism, and enhanced cellular apoptosis and senescence, accompanied by reduced DNA replication and repair. Overall, these findings imply that succinate accumulation in ADSC triggers inflammatory response and mitochondrial dysfunction, potentially contributing to a decline of cellular biological activity. Targeting succinate may offer therapeutic potential for metabolic disorders.

A surge in environmental pollution compels society to utilize food processing wastes to produce valuable compounds. Enzymatic technology, specifically cellulase-mediated hydrolysis, provides an eco-friendly and effective approach for treating food processing leftovers. The main objective of this review is to explore the significant contributions of cellulase, both in industrial settings and from an environmental perspective. Therefore, this review covers all the aspects of cellulase structural identification, classification, and evolution to its profound applications. The review initially explores cellulases’ structural and functional characteristics based on the catalytic and cellulose-binding domains and discusses cellulases’ evolutionary origin. A thorough understanding of cellulase properties is essential for overcoming the challenges associated with its commercial production for various applications. In this regard, the optimization for cellulase production through several approaches, including rational design, direct evolution, genetic engineering, and fermentation technology, is also reviewed. In addition, it also underscores the significance of agro-industrial biorefineries, which provide scalable and sustainable solutions to meet future demands for food, chemicals, materials, and fuels. Finally, the last sections of the review solely highlight the potential applications of microbial cellulases in bioremediation. In summary, this review outlines the role of cellulase in efficient valorization aimed at producing multiple bioproducts and the enhancement of environmental remediation efforts.

Exosomes are essential components produced by all cell types, originating from the endosomal pathway through the invagination of the cell membrane. Their unique physicochemical characteristics are crucial for various commercial applications. Typically, exosomes range in size from 50 to 200 nm. Exosomes derived from plant cells are larger than their animal cell counterparts and demonstrate a broader therapeutic potential. This review explores the promising research opportunities associated with plant-derived exosomes, summarizing studies on their biogenesis, characterization, isolation methods, and therapeutic applications. It also emphasizes the importance of targeted drug delivery and provides insights into engineering plant-derived exosomes with various drugs. Additionally, highlights of plant-derived exosomes as natural nano-inducers that facilitate inter-kingdom communication and cross-kingdom regulatory interactions are also elucidated herein. Henceforth, this study culminates in a multidimensional insight for innovative therapeutic strategies and biotechnological advancements in plant-derived exosome research.

Butenyl-spinosyn is a high-quality biological insecticide produced by Saccharopolyspora pogona that effectively targets a broad range of insect pests. However, the large-scale production of this insecticide is hindered by its low yield. Herein, based on prior comparative genomic analysis, five mutations were individually overexpressed in aG6. Subsequently, the combinatorial overexpression of sp1322 (encoding NAD-glutamate dehydrogenase) and sp6746 (encoding dTDP-glucose 4,6-dehydratase) in aG6 resulted in strain O1322-6746. The production of butenyl-spinosyn in O1322-6746 was 77.1% higher than that in aG6. Comparative targeted metabolomic analysis uncovered that O1322-6746 exhibited increased metabolic flux toward butenyl-spinosyn precursors. Furthermore, single-factor experiments, Plackett-Burman analysis and response surface methodology were performed to optimize the fermentation medium for O1322-6746. Ultimately, butenyl-spinosyn production was enhanced to 298.5 mg/L in a 5-L bioreactor, marking the highest yield ever reported. This work demonstrated that combining metabolic engineering with medium optimization is an effective strategy to improve butenyl-spinosyn production.