Search Results (445)

Precise regulation of micropipette outlet flow is critical for fluorescence imaging in vivo micromanipulations. In such procedures, a micropipette with a micro-sized opening is driven by gas pressure to deliver internal solution into the in vivo environment. The outlet flow rate needs to be precisely regulated to ensure a uniform and stable fluorescence distribution. However, conventional manual pressure injection methods face inherent limitations, including insufficient precision and poor reproducibility. Existing commercial microinjection systems lack a quantitative relationship between pressure and flow rate. And existing calibration methods in the field of microfluidics suffer from a limited flow-rate measurement resolution, constraining the establishment of a precise pressure–flow quantitative relationship. To address these challenges, we developed a closed-loop pressure regulation system with 1 Pa-level control resolution and established a quantitative calibration of the pressure–flow relationship using a droplet-based method. The calibration revealed a linear relationship with a mean pressure–flow gain of 4.846 $\times {10}^{-17}{\mathrm{m}}^3·{\mathrm{s}}^{-1}·{\mathrm{P}\mathrm{a}}^{-1}$ (R² > 0.99). Validation results demonstrated that the system achieved the target outlet flow rate with a flow control error less than 10 fL/s. Finally, the application results in brain-slice environment confirmed its capability to maintain stable fluorescence imaging, with fluorescence intensity fluctuations around 1.3%. These results demonstrated that the proposed approach provides stable, precise, and reproducible flow regulation under physiologically relevant conditions, thereby offering a valuable tool for in vivo micromanipulation and detection. Full article

Newborn mice (up to 6 d after birth) are suitable for genetic manipulations, such as facial vein-mediated injection, owing to their hairless and thin skin. Their small body volumes also facilitate the rapid dissemination of injected solutions, supporting gene engineering-related experiments. However, anesthesia in newborns is challenging because of the potential risks associated with anesthetic agents. Isoflurane inhalation anesthesia is an option, although its effects on brain development remain under investigation. In this study, we established a reproducible protocol for delivering nucleic acids to juvenile mouse testes using a simple isoflurane-based anesthetic system prepared from common laboratory equipment. Using this system, nucleic acids were successfully delivered to juvenile mouse testes via intra-testicular injection, followed by in vivo electroporation. The present isoflurane-based method achieved >90% postoperative survival with normal maternal nursing observations. Gene delivery resulted in limited transfection of seminiferous tubules but efficient interstitial Leydig cell transfection. Thus, gene engineering in somatic and germ cells in neonatal mice will be facilitated using the anesthetic protocol established in this study. Full article

Magnetic surgical instruments are primarily driven by magnetic force and/or micro-motors. When micro-motors are used to drive motion, they are typically installed near the manipulator joints, resulting in a larger manipulator size due to the presence of micro-motors. We designed a magnetic anchored and cable-driven surgical forceps, which separates micro-motors from the manipulator through cables. The cables are responsible for transmitting motion and force from micro-motors to the manipulator. This design enables the integration of relatively large motors (diameter: 8 mm) while maintaining a compact overall diameter of the manipulator (diameter: 10 mm). This is beneficial for improving the flexibility of the manipulator and facilitating the coordination between surgical instruments. The manipulator of the magnetic anchored and cable-driven surgical forceps has three degrees of freedom (DoFs): pitch, yaw and clamping. A magnetic attraction experiment was conducted to measure the magnetic force on the magnetic surgical forceps with the variation of abdominal skin thickness. The results indicate that at a distance of 20 mm, the magnetic force exerted on the magnetic surgical forceps is 5.86 N, with a maximum vertical load capacity of 5.13 N. Additionally, an ex vivo experiment was conducted to validate the practicality of the magnetic anchored and cable-driven surgical forceps prototype. Full article

Cell motility is a process central to life and is undoubtedly influenced by mechanical and chemical signals. Even so, other stimuli are also involved in controlling cell migration in vivo and in vitro. Among these, electric fields have been shown to provide a powerful and programmable cue to manipulate cell migration. There is now a clear consensus that the electromigration of membrane components represents the first response to an external electric field, which subsequently activates downstream signals responsible for controlling cell migration. Here, we focus on a specific mode of electrotaxis: frictionless, amoeboid-like migration. We used the Finite Element Method to solve an active gel model coupled with a mathematical model of the electromigration of aquaporins and investigate the effect of electric fields on ameboid migration. We demonstrate that an electric field can polarize aquaporins in a cell and, consequently, that the electromigration of aquaporins can be exploited to regulate water flux across the cell membrane. Our findings indicate that controlling these fluxes allows modulation of cell migration velocity, thereby reducing the cell’s migratory capacity. Our work provides a mechanistic framework to further study the impact of electrotaxis and to add new insights into specific modes by which electric fields modify cell motility. Full article

The stomach epithelium is a highly dynamic tissue that undergoes continuous self-renewal and responds robustly to injury through tightly regulated repair processes. Organoids have emerged as powerful tools for modelling gastrointestinal biology. This review focuses on the capacity of gastric organoids to model epithelial homeostasis, injury and repair in the stomach. We examine how organoid systems recapitulate key features of in vivo gastric architecture and stem cell dynamics, enabling detailed interrogation of lineage specification, proliferative hierarchies and regional identity. Gastric organoids have proven particularly useful for studying how environmental factors, such as Helicobacter pylori infection or inflammatory cytokines, disrupt epithelial equilibrium and drive metaplastic transformation. Furthermore, we discuss the emerging use of injury-mimicking conditions, co-cultures and bioengineered platforms to model regeneration and inflammatory responses in vitro. While organoids offer unparalleled accessibility and experimental manipulation, they remain limited by the absence of critical niche components such as immune, stromal and neural elements. Nevertheless, advances in multi-cellular and spatially resolved organoid models are closing this gap, making them increasingly relevant for disease modelling and regenerative medicine. Overall, gastric organoids represent a transformative approach to dissecting the cellular and molecular underpinnings of stomach homeostasis and repair. Full article

The reliability of molecular diagnostic and prognostic tools is contingent on the quality of biospecimens, which are often collected during surgical procedures. This study investigated the impact of surgical manipulation on gene expression in the urinary bladder mucosa during radical cystectomy. Seventeen patients with urinary bladder cancer were enrolled, and paired pre- and post-surgery biopsies were analyzed. Pre-surgical biopsies were obtained in situ under anesthesia, while post-surgical biopsies were collected ex vivo following bladder removal. Total RNA was extracted, and gene expression was assessed using qPCR arrays, measuring the expression of 374 inflammation-related genes. The findings from the exploratory phase were further validated by analyzing key genes in an independent patient cohort using TaqMan^® gene-specific assays. Exploratory analysis revealed significant differential expression in 27 genes, with key genes such as IL6, FOS, and PTGS2 being upregulated post-surgery. Validation of five selected genes in an independent cohort confirmed these findings. This study reinforces the necessity of accounting for surgery-induced alterations in gene expression when analyzing tissue samples collected intraoperatively. By elucidating the molecular impact of surgical interventions, this work provides critical insights for refining experimental methodologies and enhancing the interpretability of gene expression studies in clinical and research settings. Full article

Perfluorooctane sulfonate (PFOS) is a representative persistent organic pollutant that exerts toxic effects on aquatic organisms. As an alternative to PFOS, sodium p-perfluorous nonenoxybenzene sulfonate (OBS) has been frequently detected in aquatic environments and human tissues in recent years. However, its toxic effects on aquatic organisms and potential health risks to humans remain unclear. Zebrafish embryos are transparent and amenable to in vivo manipulation and observation. Therefore, in the present study, we investigated its developmental toxicity in zebrafish embryos, with PFOS as the positive control. We exposed zebrafish embryos to different concentrations of OBS (15, 20, and 25 mg/L) and PFOS (15 mg/L) for 2–168 h post fertilization (hpf) and then examined physiological and gene expression changes. At 24 hpf, spontaneous twitches in the 25 mg/L OBS group decreased to (5 ± 0.34)/min. By 48 hpf, the 20 mg/L OBS group’s hatching rate was (47.78 ± 2.22)%, significantly lower than the control. At 72 hpf, heart rates in both the PFOS and OBS groups were elevated, at 82 ± 0.6, 84.5 ± 0.5, 89.4 ± 0.3, and 93.7 ± 0.4, respectively. Similarly to PFOS, OBS induced developmental toxicity in zebrafish embryos. In addition, both OBS and PFOS exposure downregulated the expression level of anti-apoptotic Bcl-2 in zebrafish embryos, with a notable 0.53-fold decrease observed in the 25 mg/L OBS group. Conversely, they upregulated the expression levels of pro-apoptotic Bax, Caspase-3, and Caspase-9, with Caspase-3 expression increasing 1.14-, 1.5-, and 1.7-fold in the 15 mg/L PFOS, 20 mg/L OBS, and 25 mg/L OBS groups, respectively. These OBS- and PFOS-induced changes in gene expression increased apoptosis, suggesting that OBS can induce developmental toxicity in zebrafish embryos, and that its effect is comparable to that of PFOS. Therefore, considering its aquatic toxicity, measures aimed at limiting or remediating OBS pollution in the environment are necessary. Full article

Gastrointestinal (GI) cancers represent a major global health burden. Among them, colorectal cancer (CRC) is the most common type, followed by esophagus, stomach, liver, and pancreatic cancer. Since disturbance of the gut microbiota has been directly associated with the development of severe health issues, including cancer, probiotic administration may induce dysbiosis reversion and ameliorate carcinogenesis. Therefore, manipulation of the gut microbiota composition based on probiotic utilization has gradually attained scientific interest as a potent therapeutic modality for GI cancers. This review aims to synthesize the current in vitro and in vivo evidence on probiotics’ effectiveness in GI cancer chemoprevention and treatment. It also provides a classification of the fundamental anticancer features of probiotics, including antiproliferation and cell death induction, anticarcinogenic compound production, reduction in chemotherapy-related toxicity, gut microbiota modulation, intestinal barrier improvement, antioxidant activity, immunomodulatory/anti-inflammatory effects, and carcinogen detoxification. Finally, it underscores the future perspectives and challenges of probiotic administration to individuals. In this regard, it emphasizes the exploitation of advanced encapsulation techniques and the development of novel genetically engineered probiotics and next-generation probiotics as feasible ways to improve their bioavailability, ensure their targeted delivery, and eliminate their mild side effects to the host’s health. Full article

Mesenchymal stem cells (MSCs) are a type of multipotent, non-hematopoietic cells of mesodermal origin. Due to their strong immunomodulatory, immunosuppressive, and regenerative potential, MSCs are used in cell therapy for inflammatory, immune-mediated, and degenerative diseases. Exosomes derived from MSCs have several advantages over MSC therapy, including non-immunogenicity, lack of infusion toxicity, ease of isolation, manipulation, and storage, cargo specificity, and the absence of tumor-forming potential and ethical concerns. We hypothesized that preconditioning human umbilical cord blood mesenchymal stem cells (hUCB-MSCs) with the proinflammatory cytokines interleukin 17 (IL-17), IL-22, and tumor necrosis factor alpha (TNF-α), the increased levels of which are typical in psoriasis patients, can significantly increase the therapeutic efficacy of both hUCB-MSCs and their exosomes (hUCB-MSC-Exo). Our aim was to compare the therapeutic effects of hUCB-MSCs preconditioned with various combinations of proinflammatory cytokines and their hUCB-MSC-Exo, in an in vivo imiquimod-induced psoriasis-like skin inflammation model in mice. Our results showed a significant attenuation of psoriasis symptoms (erythema, scaling, and skin thickness) in mice treated with intact hUCB-MSCs, hUCB-MSCs preconditioned with IL-22 and TNF-α, and hUCB-MSC-Exo preconditioned with IL-17, IL-22 and TNF-α (MSC-Exo 3C). However, the most pronounced therapeutic effect was observed with MSC-Exo 3C treatment. In summary, we demonstrated that MSC-Exo 3C transplantation has therapeutic potential for treating psoriasis-like skin lesions. Full article

Background: The red nucleus (RN) is a phylogenetically conserved structure within the midbrain that is traditionally associated with general motor coordination; however, its specific role in controlling directional movement remains poorly understood. Methods: This study systematically investigates the function and mechanism of RN neurons in directional movement by combining stereotactic brain injections, fiber photometry recordings, multi-unit in vivo electrophysiological recordings, optogenetic manipulation, and anterograde trans-synaptic tracing. Results: We analyzed mice performing standardized T-maze turning tasks and revealed that anatomically distinct RN neuronal ensembles exhibit direction-selective activity patterns. These neurons demonstrate preferential activation during ipsilateral turning movements, with activity onset consistently occurring after movement initiation. We establish a causal relationship between RN neuronal activity and directional motor control: selective activation of RN glutamatergic neurons facilitates ipsilateral turning, whereas temporally precise inhibition significantly impairs the execution of these movements. Anterograde trans-synaptic tracing using H129 reveals that RN neurons selectively project to spinal interneuron populations responsible for ipsilateral flexion and coordinated limb movements. Conclusions: These findings offer a framework for understanding asymmetric motor control in the brain. This work redefines the RN as a specialized hub within midbrain networks that mediate lateralized movements and offers new avenues for neuromodulatory treatments for neurodegenerative and post-injury motor disorders. Full article

Recent studies have demonstrated the clinical potential of nucleic acid therapeutics (NATs). However, their efficient and scalable delivery remains a major challenge for both ex vivo and in vivo gene therapy. Microfluidic platforms have emerged as a powerful tool for overcoming these limitations by enabling precise intracellular delivery and consistent therapeutic carrier fabrication. This review examines microfluidic strategies for gene delivery at the cellular level. These strategies include mechanoporation, electroporation, and sonoporation. We also discuss the synthesis of lipid nanoparticles, polymeric particles, and extracellular vesicles for systemic administration. Unlike conventional approaches, which treat ex vivo and in vivo delivery as separate processes, this review focuses on integrated microfluidic systems that unify these functions. For example, genetic materials can be delivered to cells that secrete therapeutic extracellular vesicles (EVs), or engineered cells can be encapsulated within hydrogels for implantation. These strategies exemplify the convergence of gene delivery and carrier engineering. They create a single workflow that bridges cell-level manipulation and tissue-level targeting. By synthesizing recent technological advances, this review establishes integrated microfluidic platforms as being fundamental to the development of next-generation NAT systems that are scalable, programmable, and clinically translatable. Full article

The recent discovery of TIGR-Tas (Tandem Interspaced Guide RNA-Targeting Systems) marks a major advance in the field of genome editing, introducing a new class of compact, programmable DNA-targeting systems that function independently of traditional CRISPR-Cas pathways. TIGR-Tas effectors use a novel dual-spacer guide RNA (tigRNA) to recognize both strands of target DNA without requiring a protospacer adjacent motif (PAM). These Tas proteins introduce double-stranded DNA cuts with characteristic 8-nucleotide 3′ overhangs and are significantly smaller than Cas9, offering delivery advantages for in vivo editing. Structural analyses reveal homology to box C/D snoRNP proteins, suggesting a previously unrecognized evolutionary lineage of RNA-guided nucleases. This review positions TIGR-Tas at the forefront of a new wave of RNA-programmable genome-editing technologies. In parallel, I provide comparative insight into the diverse and increasingly modular CRISPR-Cas systems, including Cas9, Cas12, Cas13, and emerging effectors like Cas3, Cas10, CasΦ, and Cas14. While the CRISPR-Cas universe has revolutionized molecular biology, TIGR-Tas systems open a complementary and potentially more versatile path for programmable genome manipulation. I discuss mechanistic distinctions, evolutionary implications, and potential applications in human cells, synthetic biology, and therapeutic genome engineering. Full article

Livestock farming is a vital component of global food security, yet it remains a major contributor to greenhouse gas (GHG) emissions, particularly methane (CH₄), which has a global warming potential 28 times greater than carbon dioxide (CO₂). This review provides a comprehensive synthesis of current knowledge surrounding the sources, biological mechanisms, and mitigation strategies related to CH₄ emissions from ruminant livestock. We first explore the process of methanogenesis within the rumen, detailing the role of methanogenic archaea and the environmental factors influencing CH₄ production. A thorough assessment of both direct and indirect methods used to quantify CH₄ emissions is presented, including in vitro techniques (e.g., syringe method, batch culture, RUSITEC), in vivo techniques (e.g., respiration chambers, Greenfeed, laser CH₄ detectors), and statistical modeling approaches. The advantages and limitations of each method are critically analyzed in terms of accuracy, cost, feasibility, and applicability to different farming systems. We then examine a wide range of mitigation strategies, organized into four core pillars: (1) animal and feed management (e.g., genetic selection, pasture quality improvement), (2) diet formulation (e.g., feed additives such as oils, tannins, saponins, and seaweed), (3) rumen manipulation (e.g., probiotics, ionophores, defaunation, vaccination), and (4) manure management practices and policy-level interventions. These strategies are evaluated not only for their environmental impact but also for their economic and practical viability in diverse livestock systems. By integrating technological innovations with sustainable agricultural practices, this review highlights pathways to reduce CH₄ emissions while maintaining animal productivity. It aims to support decision-makers, researchers, and livestock producers in the global effort to transition toward climate-smart, low-emission livestock farming. Full article

Organoids are three-dimensional tissue culture models derived from stem cells, and they have become one of the most valuable tools in biomedical research. These self-organizing miniature organs mimic the structure−function properties of their in vivo counterparts and offer an exceptional prospective for disease modeling, drug discovery, and regenerative medicine. By replicating the complexity of human tissue, organoids enable the study of disease pathophysiology, tissue development, and cellular interactions in a highly controlled and manipulable environment. Recent developments in organoid technology have enabled the production of functional organoids of various tissues. These systems have proven to be highly promising tools for personalized medicine. In addition, organoids have also raised hopes for the development of functional transplantable organs, transforming the study of regenerative medicine. This review provides an overview of the current state of organoid technology and its application and prospects and focuses on the transformative impact of organoid technology on biomedical research and its contribution to human health. Full article

Stress granules (SG), dynamic cytoplasmic condensates formed via liquid-liquid phase separation (LLPS), serve as a critical hub for cellular stress adaptation and antiviral defense. By halting non-essential translation and sequestering viral RNA, SG restrict viral replication through multiple mechanisms, including PKR-eIF2α signaling, recruitment of antiviral proteins, and spatial isolation of viral components. However, viruses have evolved sophisticated strategies to subvert SG-mediated defenses, including proteolytic cleavage of SG nucleators, sequestration of core proteins into viral replication complexes, and modulation of stress-responsive pathways. This review highlights the dual roles of SG as both antiviral sentinels and targets of viral manipulation, emphasizing their interplay with innate immunity, autophagy, and apoptosis. Furthermore, viruses exploit SG heterogeneity and crosstalk with RNA granules like processing bodies (P-bodies, PB) to evade host defenses, while viral inclusion bodies (IBs) recruit SG components to create proviral microenvironments. Future research directions include elucidating spatiotemporal SG dynamics in vivo, dissecting compositional heterogeneity, and leveraging advanced technologies to unravel context-specific host-pathogen conflicts. This review about viruses and SG formation helps better understand the virus-host interaction and game process to develop new drug targets. Understanding these mechanisms not only advances virology but also informs innovative strategies to address immune escape mechanisms in viral infections. Full article