Search Results (1,019)

Amplex Red is a widely used fluorogenic probe for hydrogen peroxide (H₂O₂) detection. In the presence of horseradish peroxidase (HRP), it reacts with H₂O₂ to form fluorescent resorufin. However, during high-voltage electric pulse treatment, fluorescence readouts may be influenced not only by H₂O₂ generation but also by the medium composition, pulse conditions, pH-dependent effects, and metal ions released as a result of electrode corrosion. In this study, we examined how medium composition and selected metal ions relevant to electroporation conditions, such as Fe²⁺, Fe³⁺, Al³⁺, and Cr⁶⁺, affect the Amplex Red/HRP/H₂O₂ fluorescence signal. Fluorescence intensity differed markedly among media, with the highest signal in phosphate-buffered saline (PBS, ~34,000 a.u.), followed by HB1 buffer (~19,000 a.u.), cell culture medium (~9000 a.u.), and distilled water (~4100 a.u.). To model ion-mediated interference, metal ions were added directly to the assay mixture. All tested ions reduced fluorescence in a concentration-dependent manner. At 0.4 µM H₂O₂, Fe²⁺ reduced fluorescence by up to 92%, Fe³⁺ by 60%, Al³⁺ by 35%, and Cr⁶⁺ by 40%. At 10 µM H₂O₂, the quenching effect was reduced but remained evident. In addition, the fluorescence readout showed strong dependence on medium composition and its pH, further demonstrating its vulnerability to the chemical environment. These findings show that the Amplex Red assay is highly sensitive to the assay environment and metal-ion interference. Therefore, it should not be used as a standalone, interference-free measure of H₂O₂ in electroporation-related experiments without appropriate validation controls. Full article

Galacto-oligosaccharides (GOS) are widely recognized as prebiotic ingredients, whereas their technological role in stabilizing lipid-based systems remains poorly understood. In this study, GOS were synthesized by enzymatic transgalactosylation of lactose and enriched by ethanol precipitation. Carbohydrate composition, glass transition temperature (Tg), and their influence on the formation of primary lipid oxidation were evaluated using freeze-dried lecithin liposomes as a model membrane system. Ethanol precipitation increased the GOS content from 22.8% to 50.7% (w/w) while reducing the proportion of monosaccharides from 40.0% to 15.5% (w/w). Tg was strongly influenced by carbohydrate composition and relative humidity (RH), with GOS-enriched preparations exhibiting higher Tg values than the crude reaction mixture. The formation of primary lipid oxidation products, assessed by UV spectroscopy through conjugated diene formation at 234 nm, was significantly lower in liposomes freeze-dried with GOS-rich preparations or sucrose than in phosphate-buffered saline (PBS) controls, particularly under storage conditions in which the carbohydrate matrices remained in the glassy state. At higher RH, transition to the rubbery state was associated with increased conjugated diene formation. These findings suggest that the protective effect of GOS-rich formulations is associated with the physicochemical properties of the carbohydrate matrix and is consistent with the proposed role of the glassy state in limiting molecular mobility, rather than direct antioxidant activity. Beyond their established prebiotic properties, GOS-rich preparations may also contribute to the stability of dehydrated lipid-containing systems, supporting their use as multifunctional food ingredients. Full article

Oral mucositis is a frequent adverse effect of cancer therapy and is often complicated by pain, oral dryness, and secondary infection. Freeze-dried hyaluronic acid (HA) sheets can protect and moisturize the oral mucosa, but their intrinsic antibacterial activity is limited. We developed freeze-dried HA sheets containing the commercial persimmon tannin product PANCIL^® PS-M and evaluated their release behavior, pH profile, antimicrobial activity, and in vitro cytocompatibility. Persimmon tannin release into phosphate-buffered saline was quantified over 8 h using the Folin–Ciocalteu method. Antimicrobial activity against Streptococcus mitis, Prevotella intermedia, and Candida albicans was assessed by colony counting, and cytocompatibility was evaluated using the human oral cell line T3M-1. Approximately 95% of the incorporated persimmon tannin was released within 8 h, while the surrounding pH remained near neutral to mildly alkaline. The antibacterial HA sheets markedly reduced viable counts of S. mitis and P. intermedia, whereas no inhibitory effect was observed against C. albicans. Neither HA sheet nor antibacterial HA sheet solutions significantly reduced T3M-1 cell viability at concentrations up to 100 µg/mL after 24 h. These findings suggest that persimmon tannin-containing HA sheets may provide sustained antibacterial activity without detectable cytotoxicity under the tested conditions and may serve as a topical adjunct for cancer treatment-related oral mucositis. Full article

Corneal endothelium is a single layer of polygonal cells responsible for maintaining stromal dehydration and corneal transparency. Isolating corneal endothelium from murine models is exceptionally challenging without an established protocol, which historically limits the utility of mouse models to study corneal endothelial disease. Herein, a detailed method is presented to obtain a highly pure population of corneal endothelial cells via mechanical peeling of the endothelial sheet from both young and aged mice. Under a dissecting microscope, the corneal dome, including the limbus, was dissected. Following the removal of the iris and trabecular meshwork, the corneal dome was rinsed in phosphate-buffered saline (PBS). Fine, blunt forceps were utilized to stabilize the edge of the corneal dome, while ultra-fine forceps were employed to grasp the edge of Descemet’s membrane (DM). The edge of the DM was identified by performing a gentle micro-scratch along the limbal margin until a small flap emerged. Subsequently, the entire endothelial sheet along with the DM was carefully peeled from the periphery toward the corneal center. The isolated endothelial sheet can be dried flat on a microscope slide to investigate spatial gene expression, cell morphology, and cellular interactions. To obtain a single-cell suspension, the endothelial sheet was enzymatically digested in 0.25% Trypsin-EDTA for 8 min at 37 °C, yielding a highly purified population of corneal endothelial cells (95.7%). This protocol demonstrated high reproducibility for both young (approximately 1 month old) and aged (approximately 1 year old) mouse corneas. The total cell yields collected from young and old mice were 3634 and 4560 cells per eye, respectively, exhibiting high cell viability (young: 98.3%, old: 95.8%) and a high percentage of single cells (young: 90.6%, old: 90.1%). This detailed isolation protocol will facilitate advanced downstream research on the corneal endothelium using murine models. Full article

Background/Objectives: Psoriasis is a chronic, immune-mediated inflammatory skin disorder affecting millions of individuals worldwide. It remains a therapeutic challenge due to the limited skin penetration of many drugs, adverse systemic effects, and the need for long-term management. Transferosomal nanoformulations containing natural products were proposed as a potential therapeutic tool. Methods: Transferosomes containing 20-hydroxyecdysone and resveratrol were prepared using the thin-film hydration method followed by probe ultrasonication, generating several formulation variants differing in composition. The vesicles were characterized by dynamic light scattering (DLS) and nanoparticle tracking analysis (NTA) to determine size, polydispersity index (PDI), and zeta potential. Furthermore, time-domain nuclear magnetic resonance (TD-NMR) relaxation measurements were employed to evaluate local molecular dynamics and membrane fluidity, providing deeper insights into the structural integrity and elasticity of the transferosomal systems. The stability of the nanoformulations was assessed for one month in water and phosphate-buffered saline (PBS). Viability was evaluated in vitro using the MTS assay on human epidermal keratinocyte (HEK) and psoriasis-patient derived human epidermal keratinocyte (PHEK) cell lines, alongside antimicrobial profiling against four representative human skin microbiome strains. Acute toxicity was further examined in vivo using the Danio rerio FET test. Results: The formulations demonstrated high physicochemical stability over the tested period, maintained desirable particle sizes within 100–200 nm, and showed no cytotoxicity toward skin-associated bacteria or in the zebrafish model. Conclusions: The developed nanoformulations present suitable properties in terms of safety and stability and can be considered for potential use in psoriasis treatment. Full article

Background: Patients with Kawasaki disease (KD) who develop coronary artery aneurysms (CAAs) are at increased risk of future fatal coronary events. Pharmacotherapeutic strategies to prevent coronary stenosis are still lacking. In this exploratory study, the therapeutic effect of the angiotensin receptor blocker (ARB) losartan on coronary artery (CA) stenosis was investigated in a murine model. Methods: Five-week-old male C57BL/6J mice were intraperitoneally injected with 1000 μg of Lactobacillus casei cell wall extract (LCWE) (n = 12) to induce CA stenosis. Two weeks later, the LCWE-injected mice (n = 12) were divided into two groups: six received drinking water containing losartan (100 mg/L) (LCWE+ARB), while six received normal drinking water (LCWE group). A control group (n = 5) received phosphate-buffered saline (PBS) instead of LCWE. Sixteen weeks after LCWE administration—corresponding to the peak of CA stenosis and 14 weeks after treatment initiation—the mice were euthanized for histological evaluation of the coronary arteries. Results: Losartan treatment significantly reduced the coronary arteritis score (median [IQR (interquartile range)]: 0 [0–9.5] vs. 21.5 [15–24.3], p = 0.003). LCWE-induced neointimal formation with vascular smooth muscle cell proliferation and subsequent CA stenosis were markedly attenuated in losartan-treated mice (25% vs. 100%, p < 0.001). Losartan-associated attenuation of CA stenosis was accompanied by the preservation of medial calponin expression, a reduction in the number of proliferating cell nuclear antigen (PCNA)-positive cells in the neointima, and a decrease in serum MMP-9 (matrix metalloproteinase-9) levels. Conclusions: These findings from a murine model of KD provide preliminary evidence that losartan may attenuate coronary artery remodeling. Further mechanistic studies are warranted to clarify its potential translational relevance. Full article

Background: Buffaloes have a significant share in the agricultural economies of many developing countries; however, the industry is not well-established because of the low reproductive efficiency of buffaloes. In this study, we designed a novel strategy to enhance buffalo reproductive efficiency and examined the significance of intramuscular AMH-INH-RFRP DNA vaccine immunization on reproductive performance. Methods: Allocated into four groups (n = 30/group), female buffaloes (aged 5–8 years, n = 120) were randomly enrolled in the study. Treatment groups received 10 mL of AMH-INH-RFRP DNA vaccine intramuscularly, once daily for three consecutive days, at the following concentrations: T1—3 × 10⁸ CFU/mL, T2—3 × 10⁹ CFU/mL, and T3—3 × 10¹⁰ CFU/mL. All vaccinated groups received a booster vaccination two weeks later, while the control group received 10 mL of phosphate-buffered saline (PBS) intramuscularly on the same schedule. Serum antibody titers against anti-Müllerian hormone (AMH), inhibin (INH), and RF-amide-related peptide (RFRP) were quantified using indirect ELISA on days 14 (post-primary immunization) and 28 (post-booster immunization). Results: Following primary immunization, antibody titers against AMH, INH, and RFRP were significantly higher in the T3 group than in the T1 group (p < 0.05), and booster immunization further increased antibody positivity rates. Serum concentrations of IL-4, IFN-γ, and E₂ were significantly elevated in the T2 and T3 groups compared to the control group (p < 0.05), while P₄ levels showed no significant differences among groups. Ultrasonography revealed that the ovulatory follicle diameter and dominant follicle growth rate were significantly increased in the T2 and T3 groups compared to the control and T1 groups (p < 0.05). The estrus rate was significantly higher in the T3 group than in the control group (76.67% vs. 40.00%, p < 0.05), and ovulation rates were significantly higher in the T2 (83.33%) and T3 (86.67%) groups compared to the control group (56.67%) (p < 0.05). However, conception rates did not differ significantly between vaccinated and control groups in the primary dose comparison (p > 0.05). In exploratory post hoc analyses, antibody-positive (Ab+) buffaloes exhibited higher E₂ concentrations, larger ovulatory follicle diameters, accelerated dominant follicle growth rates, and higher estrus and ovulation rates compared to antibody-negative (Ab−) buffaloes. Furthermore, gestating buffaloes were monitored until calving, revealing no significant effect of the vaccine on newborn calf weight or body size. Conclusions: In total, intramuscular immunization with the AMH-INH-RFRP DNA vaccine, particularly at the highest dose, enhances reproductive performance in buffaloes by stimulating E₂ secretion and follicular development. Full article

The growing market for insect-based food and feed ingredients raises concerns about authenticity, labelling transparency, and regulatory compliance, highlighting the need for rapid screening methods capable of distinguishing selected insect-derived materials. In this preliminary feasibility study, Raman spectroscopy combined with partial least squares discriminant analysis (PLS-DA) was evaluated for the fingerprinting and differentiation of PBS-extractable components from nine insect species. Extracts of 96 dried insect samples prepared in phosphate-buffered saline (PBS, pH 9.4) were analysed, yielding 358 insect spectra after quality control; PBS spectra were included as a technical control class. The overall correct classification rate (CCR) reached 61.6%, indicating moderate classification performance rather than validated species-level authentication. Apis mellifera drones achieved 100.0% CCR, while Hermetia illucens and Tenebrio molitor both reached 80.0%. Complete misclassification of Shelfordella tartara as Blaptica dubia indicated strong spectral similarity between closely related cockroach species. VIP score analysis identified the 1700–1800 cm⁻¹ region, corresponding primarily to lipid ester carbonyl vibrations, as the most discriminatory spectral range. The results suggest that Raman spectroscopy can provide taxon-related biochemical fingerprints of PBS-extractable insect components, while also highlighting the need for expanded reference libraries, improved sampling strategies, and validation in complex food matrices to further develop its potential as a screening approach. Full article

Porcine circovirus type 2 (PCV2) and Mycoplasma hyopneumoniae (Mhp) are major contributors to the porcine respiratory disease complex. Although PCV2-Mhp bivalent inactivated vaccines are useful for simultaneous disease control, their immunogenicity may be improved by adjuvant optimization. This study evaluated a composite adjuvant consisting of porcine interleukin-2 (IL-2), IL-4, IL-6 and MnJ(beta), a colloidal manganese adjuvant, in an initial murine immunogenicity model. Thirty female Kunming mice were assigned to three groups (n = 10/group): bivalent antigen plus IL-2/IL-4/IL-6/MnJ(beta), bivalent antigen plus MnJ(beta), or phosphate-buffered saline. Body weight, complete blood count, peripheral blood T- and B-cell subsets, PCV2-specific IgG and Mhp-specific indirect hemagglutination titers were monitored after primary and booster immunization. The composite formulation did not suppress body-weight gain or induce sustained abnormalities in erythrocyte- or platelet-related indices. WBC, neutrophil, lymphocyte and monocyte counts were elevated in group A at days 7 and 28 post-primary immunization, indicating transient immune activation. Day-56 flow cytometry indicated increased CD19+IgM-IgD- B-cell and effector/memory T-cell-associated responses. PCV2-specific IgG increased from day 14 onward. At day 56, the OD450 value in group A reached 1.532 ± 0.006, compared with 1.095 ± 0.004 in group C1 and 0.102 ± 0.002 in group C2, corresponding to approximately 1.40-fold and 15.09-fold higher levels than the MnJ(beta)-adjuvanted and PBS control groups, respectively. Mhp-specific IHA titers were also maintained at high levels after booster immunization; at day 56, group A showed a log2 endpoint titer of 13.00 ± 0.00, corresponding to a GMT of 1:8192, whereas group C1 showed a log2 endpoint titer of 12.00 ± 0.00, corresponding to a GMT of 1:4096, and group C2 remained negative. These results indicate that the IL-2/IL-4/IL-6/MnJ(beta) composite adjuvant demonstrates potential for improving antibody and peripheral lymphocyte responses to PCV2-Mhp bivalent antigen, but protective efficacy must be confirmed in target-species challenge studies. Full article

In this study, we assessed the efficacy of a single intra-articular injection of parathyroid hormone (PTH) (1–34) to enhance the meniscal healing of a medial meniscal radial tear. A radial tear involving two-thirds of the meniscal width was created in the anterior medial meniscus of the bilateral knees in 30 Japanese white rabbits that were injected with either PTH (1–34) or phosphate-buffered saline. Macroscopic, biomechanical, and histological assessments of the meniscus were performed at 4, 8, and 12 weeks post-surgery. PTH injection resulted in an improved meniscal healing rate at 4 and 8 weeks compared with phosphate-buffered saline (p = 0.05 and p < 0.05, respectively). After 4 weeks, PTH injection led to a higher circumferential strength in biomechanical testing. The meniscal healing score exhibited a trend toward improvement, reaching borderline statistical significance at 4 weeks with PTH injection (1.6 ± 1.3) than with phosphate-buffered saline (0.5 ± 0.8) (p = 0.05). In the outer two-thirds of the meniscal width, picrosirius red staining revealed greater type I collagen content following PTH injection. A single injection of PTH (1–34) into the knee joint improved meniscal healing in a rabbit medial meniscus radial tear model at 4 and 8 weeks, which was accompanied by increased type I collagen content around the healing site. Full article

Background/Objectives: One-piece (monoblock) dental implants are increasingly used, particularly in patients with limited bone availability. Prosthetic alignment is often achieved via plastic deformation of the titanium implant. This study aimed to evaluate the impact of such deformation-induced residual stress on the corrosion resistance of these implants. Methods: Two types of monoblock dental implants (spherical “S” and Mag-Conical “M”) were subjected to controlled plastic deformation. Residual stress was quantified by X-ray diffraction using the Bragg–Brentano method. Electrochemical behavior was evaluated by measuring the open-circuit potential (E_OCP) and performing potentiodynamic polarization tests in phosphate-buffered saline (PBS) at 37 °C. Metal ion release (Ti, V, Al) was quantified by inductively coupled plasma mass spectrometry (ICP-MS) at specific immersion time points. Surface morphology and corrosion features were examined by scanning electron microscopy (SEM). Results: Residual stress values increased significantly after plastic deformation. The open-circuit potential (E_OCP) shifted towards more electronegative values in both implant designs as deformation-induced residual stresses increased. The E_OCP values shifted from −0.099 V to −0.227 V in the S design and from −0.115 V to −0.141 V in the M design, comparing the as-received condition with the deformed state, respectively. Potentiodynamic tests showed an increase in corrosion rate from 0.0021 mm/year for the original implants to 0.0156 mm/year for the deformed ones. Surfaces in the stressed regions exhibited a high density of corrosion pits, indicating localized electrochemical degradation. Deformed dental implants also exhibited higher ion release, particularly of titanium and vanadium, with higher levels observed in implants with greater residual stress and lower corrosion resistance. In the deformed regions, the release of titanium and vanadium ions into the surrounding medium was nearly five-fold higher. Conclusions: Plastic deformation of monoblock dental implants is associated with reduced corrosion resistance. Increased residual stress correlates with enhanced electrochemical degradation and ion release, which may have relevant implications for implant selection and clinical placement. Full article

Bone marrow aspirate concentrate (BMAC) is widely used as a source of mesenchymal stromal/stem cells (MSCs) for musculoskeletal regeneration; however, BMAC lacks essential bone marrow extracellular matrix (ECM) components, a critical component of the stem cell niche that regulates MSC survival, paracrine signaling, and regenerative capacity. We previously demonstrated that an ECM-retaining micronized bone marrow product (BMAX™) preserves pro-regenerative MSC phenotypes in vitro. Human bone marrow from a single donor was processed into conventional BMAC or BMAX™. Post-traumatic osteoarthritis was induced in immunodeficient rats using destabilization of the medial meniscus (DMM). Four weeks after surgery, animals were randomly assigned to receive intra-articular injections of BMAX™, BMAC, or phosphate-buffered saline (n = 10–12 in each group). Pain-related behavior (n = 5–6/group), histological assessment (n = 3–6/group), and micro-computed tomography (n = 4–6/group) were evaluated for up to 8 weeks after treatment. At 4 weeks after treatment, BMAC significantly increased the paw withdrawal threshold compared with PBS (p = 0.0068), whereas BMAX™ significantly reduced knee joint swelling compared with both PBS (p = 0.0235) and BMAC (p = 0.0039), and BMAX™ significantly improved knee bend scores compared with PBS (p = 0.0011). Neither treatment significantly improved OARSI histological scores at this time point. At 8 weeks after treatment, BMAX™ significantly increased the paw withdrawal threshold compared with PBS (p = 0.0305), whereas BMAC showed a non-significant trend (p = 0.0517); both treatments significantly reduced knee bend scores compared with PBS (p = 0.0027 and p = 0.0255), and BMAX™ demonstrated significantly lower knee bend scores than BMAC (p = 0.0090). BMAX™ significantly reduced knee swelling compared with PBS (p = 0.0196). Histologically, BMAX™ significantly improved OARSI scores in both the femoral condyle and tibial plateau compared with PBS (p = 0.0020 and p = 0.0003, respectively), whereas BMAC significantly improved only tibial plateau OARSI scores (p = 0.0014). Furthermore, BMAX™ demonstrated significantly lower femoral condyle OARSI scores than BMAC (p = 0.0243). Micro-computed tomography revealed that both BMAX™ and BMAC significantly reduced medial subchondral trabecular separation compared with PBS (p = 0.0340 and p = 0.0426, respectively), whereas no significant differences were observed between the two treatment groups for other bone structural parameters. In conclusion, preservation of the native bone marrow ECM was associated with improved functional outcomes and greater cartilage regeneration compared with conventional BMAC in this preclinical rat model of post-traumatic osteoarthritis. These findings support the concept that maintaining the native stem cell niche may enhance the therapeutic potential of bone marrow-derived cell therapies for osteoarthritis. Full article

The development of bioactive surgical sutures capable of delivering therapeutic agents directly at the wound site has gained increasing attention in biomedical research. Functionalized sutures may provide localized antimicrobial or anti-inflammatory activity, potentially reducing postoperative complications and promoting tissue healing. In this study, absorbable surgical sutures were impregnated with carvacrol, a natural phenolic compound with well-known antimicrobial properties, using supercritical carbon dioxide (scCO₂) technology. The impregnation process was carried out at 35 °C and 10 MPa for 120 min, allowing for the incorporation of carvacrol into the polymeric matrix of the sutures. The in vitro release behavior of the impregnated sutures was evaluated in phosphate-buffered saline (PBS, pH 7.4) at 37 °C over an 8-day period. The concentration of released compounds was determined by UV–Vis spectrophotometry using previously established calibration curves. An analysis of the experimental release data demonstrated that both carvacrol and benzydamine hydrochloride (HCl) (Tantum Verde^®) exhibited a sustained release profile throughout the incubation period. Carvacrol release increased progressively from 2.02 ± 0.15 ppm on day 1 to 7.45 ± 0.15 ppm on day 7, followed by a slight stabilization on day 8 (7.25 ± 0.31 ppm). Similarly, benzydamine HCl (Tantum Verde^®) release increased from 1.83 ± 0.11 ppm on day 1 to 3.29 ± 0.13 ppm on day 8. Release kinetics were analyzed using the Korsmeyer–Peppas model, indicating that the release mechanism was predominantly diffusion-controlled during the initial stage of the experiment. The results demonstrate that supercritical CO₂ impregnation is an effective solvent-free technique for incorporating bioactive compounds into absorbable sutures, enabling controlled release under physiological conditions. Full article

Chronic, non-healing foot ulcers are the most common complication of diabetes and are associated with high morbidity, mortality, and substantial health-care costs. Impaired wound healing in diabetes results from a complex pathophysiology involving persistent inflammation, interrupted re-epithelialization, impaired neovascularization, and defective granulation tissue formation. Reconstituted high-density lipoprotein (rHDL), with its antioxidant and anti-inflammatory properties, may counteract these processes and thereby promote wound repair. We investigated the effect of topical administration of rHDL in full thickness excisional wounds in murine and porcine diabetic models. rHDL significantly enhanced wound closure in diabetic mice in a dose-dependent manner as compared to phosphate-buffered saline (PBS) control. Histological and immunohisto-chemical analysis showed that rHDL-treated wounds had increased collagen deposition, a higher number of alpha-smooth muscle actin-positive cells, greater macrophage infiltration, and decreased neutrophil infiltration compared to PBS controls. In contrast, in the porcine model no wound healing improvement was observed after daily topical application of rHDL. Administration of rHDL enhances wound closure in a murine model of diabetic wound healing by promoting collagen production and modulating inflammation. However, lack of efficacy in a more physiologically relevant pre-clinical porcine model under the experimental conditions tested does not support further development of a topical rHDL formulation for diabetic wound healing indications. Full article

Microrobots navigating the human body require biocompatible actuators capable of functioning in physiological fluids. The microalga Euglena gracilis offers precise phototactic control; however, its operational stability in simulated physiological environments remains unproven. Here, we report that a stepwise acclimation process preserves the robotic functionality of E. gracilis in 100% phosphate-buffered saline (PBS), 100% fetal bovine serum (FBS), and a NaCl solution at a concentration equivalent to PBS (137 mM). We compared direct transfer against a graduated adaptation protocol, evaluating morphology, swimming speed, motility rate, and phototaxis. Direct transfer to each medium caused near-total immobilization, whereas stepwise acclimation retained motility. Acclimated cells exhibited size reduction (miniaturization) while maintaining their characteristic eccentricity. These adapted cells sustained a negative phototactic response among the remaining motile population, supporting optical controllability despite reduced swimming speed. These results indicate that stepwise acclimation allows E. gracilis to retain partial motility and phototactic controllability under simulated physiological saline conditions, and that the observed miniaturization and preserved photo-responsiveness may be useful features for future biohybrid microrobotics. Full article