Hypertonicity during a rapid rise in D-glucose mediates first-phase insulin secretion.

Introduction: Biphasic insulin secretion is an intrinsic characteristic of the pancreatic islet and has clinical relevance due to the loss of first-phase in patients with Type 2 diabetes. As it has long been shown that first-phase insulin secretion only occurs in response to rapid changes in glucose, we tested the hypothesis that islet response to an increase in glucose is a combination of metabolism plus an osmotic effect where hypertonicity is driving first-phase insulin secretion. Methods: Experiments were performed using perifusion analysis of rat, mouse, and human islets. Insulin secretion rate (ISR) and other parameters associated with its regulation were measured in response to combinations of D-glucose and membrane-impermeable carbohydrates (L-glucose or mannitol) designed to dissect the effect of hypertonicity from that of glucose metabolism. Results: Remarkably, the appearance of first-phase responses was wholly dependent on changes in tonicity: no first-phase in NAD(P)H, cytosolic calcium, cAMP secretion rate (cAMP SR), or ISR was observed when increased D-glucose concentration was counterbalanced by decreases in membrane-impermeable carbohydrates. When D-glucose was greater than 8 mM, rapid increases in L-glucose without any change in D-glucose resulted in first-phase responses in all measured parameters that were kinetically similar to D-glucose. First-phase ISR was completely abolished by H89 (a non-specific inhibitor of protein kinases) without affecting first-phase calcium response. Defining first-phase ISR as the difference between glucose-stimulated ISR with and without a change in hypertonicity, the peak of first-phase ISR occurred after second-phase ISR had reached steady state, consistent with the well-established glucose-dependency of mechanisms that potentiate glucose-stimulated ISR. Discussion: The data collected in this study suggests a new model of glucose-stimulated biphasic ISR where first-phase ISR derives from (and after) a transitory amplification of second-phase ISR and driven by hypertonicity-induced rise in H89-inhibitable kinases likely driven by first-phase responses in cAMP, calcium, or a combination of both.

Population-specific responses to pollution exposure suggest local adaptation of invasive red swamp crayfish Procambarus clarkii along the Mediterranean French coastline.

Anthropogenic stressors can have an impact in a broad range of physiological processes and can be a major selective force leading to rapid evolution and local population adaptation. In this study, three populations of the invasive crayfish Procambarus clarkii were investigated. They are geographically separated for at least 20 years, and live in different abiotic environments: a freshwater inland lake (Salagou lake) with no major anthropogenic influence and two other coastal wetlands regularly polluted by pesticides along the Mediterranean coast (Camargue region and Bages-Sigean lagoon). Collected adults were genetically characterized using the mitochondrial COI gene and haplotype frequencies were analyzed for genetic variability within and between populations. Results revealed a higher genetic diversity for these invasive populations than any previous report in France, with more than seven different haplotypes in a single population. The contrasting genetic diversity between the Camargue and the other two populations suggest different times and sources of introduction. To identify differences in key physiological responses between these populations, individuals from each population were maintained in controlled conditions. Data on oxygen consumption rates indicate that the Salagou and Bages-Sigean populations possess a high inter-individual variability compared to the Camargue population. The low individual variability of oxygen consumption and low genetic diversity suggest a specific local adaptation for the Camargue population. Population-specific responses were identified when individuals were exposed to a pesticide cocktail containing azoxystrobin and oxadiazon at sublethal concentrations. The Salagou population was the only one with altered hydro-osmotic balance due to pollutant exposure and a change in protease activity in the hepatopancreas. These results revealed different phenotypic responses suggesting local adaptations at the population level.

Bedside ultrasound-assisted puncture and drainage under local anesthesia: A novel approach for early post-operative space-occupying tumor bed cysts of glioma resection.

OBJECTIVES: To investigate the causes of space-occupying tumor bed cysts formed early after glioma resection by measuring the osmotic pressure gradient between cystic fluid, serum, and cerebrospinal fluid (CSF) and propose a new method of bedside ultrasound-assisted puncture and drainage (UAP&D) under local anesthesia for treatment. METHODS: Bedside UAP&D under local anesthesia was performed through a burr hole on the skull flap.Following a successful puncture, cystic fluid was collected, while blood and CSF were obtained through vein and lumbar puncture, respectively. The osmotic pressure of all fluids collected was measured. The appearance, biochemical composition, and results of microbial culture of cystic fluid and CSF were analyzed. Within 24 h after UAP&D, a CT examination and Glasgow coma scale (GCS) were assessed. RESULTS: The osmotic pressure of cystic fluid was higher than that of serum and CSF. White blood cell count and protein concentration were higher in the cystic fluid compared to the CSF. Conversely, the concentration of chloride ions and glucose were lower. CT scan confirmed the correct placement of the cysts' drainage tube and that the cysts' volume decreased significantly with continued drainage. Accompanied by a reduction in the volume of cysts, there were significant improvements in GCS score within 24 h after UAP&D. All drainage tubes were removed within 2-5 days, and no puncture tract hemorrhage or infection was observed. CONCLUSION: The osmotic pressure gradient between cystic fluid, serum, and CSF caused the formation of early post-operative space-occupying tumor bed cysts for glioma. UAP&D aligns with the concept that micro-invasive neurosurgery is an effective treatment method for such cysts.

Lumen expansion is initially driven by apical actin polymerization followed by osmotic pressure in a human epiblast model.

Post-implantation, the pluripotent epiblast in a human embryo forms a central lumen, paving the way for gastrulation. Osmotic pressure gradients are considered the drivers of lumen expansion across development, but their role in human epiblasts is unknown. Here, we study lumenogenesis in a pluripotent-stem-cell-based epiblast model using engineered hydrogels. We find that leaky junctions prevent osmotic pressure gradients in early epiblasts and, instead, forces from apical actin polymerization drive lumen expansion. Once the lumen reaches a radius of ∼12 µm, tight junctions mature, and osmotic pressure gradients develop to drive further growth. Computational modeling indicates that apical actin polymerization into a stiff network mediates initial lumen expansion and predicts a transition to pressure-driven growth in larger epiblasts to avoid buckling. Human epiblasts show transcriptional signatures consistent with these mechanisms. Thus, actin polymerization drives lumen expansion in the human epiblast and may serve as a general mechanism of early lumenogenesis.

Efficacy of different bioactive glass S53P4 formulations in biofilm eradication and the impact of pH and osmotic pressure.

AIM: The challenging properties of biofilm-associated infections and the rise of multidrug-resistant bacteria are prompting the exploration of alternative treatment options. This study investigates the efficacy of different bioactive glass (BAG) formulations - alone or combined with vancomycin - to eradicate biofilm. Further, we study the influence of BAG on pH and osmotic pressure as important factors limiting bacterial growth. METHOD: Different BAG S53P4 formulations were used for this study, including (a) powder (<45 µm), (b) granules (500-800 µm), (c) a cone-shaped scaffold and (d) two putty formulations containing granules with no powder (putty A) or with additional powder (putty B) bound together by a synthetic binder. Inert glass beads (1.0-1.3 mm) were included as control. All formulations were tested in a concentration of 1750 mg/ml in Müller-Hinton-Broth against methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant Staphylococcus epidermidis (MRSE). Vancomycin was tested at the minimum-inhibitory concentration for each strain. Changes in pH and osmolality over time were assessed at 0 h, 24 h, 72 h and 168 h. RESULTS: All tested BAG formulations showed antibiofilm activity against MRSA and MRSE. Powder and putty B were the most effective formulations suppressing biofilm leading to its complete eradication after up to 168 h of co-incubation, followed by granules, scaffold and putty A. In general, MRSE appeared to be more susceptible to bioactive glass compared to MRSA. The addition of vancomycin had no substantial impact on biofilm eradication. We observed a positive correlation between a higher pH and higher antibiofilm activity. CONCLUSIONS: BAG S53P4 has demonstrated efficient biofilm antibiofilm activity against MRSA and MRSE, especially in powder-containing formulations, resulting in complete eradication of biofilm. Our data indicate neither remarkable increase nor decrease in antimicrobial efficacy with addition of vancomycin. Moreover, high pH appears to have a direct antimicrobial impact; the role of high osmolality needs further investigation.

Sourcebook update: RBC hemolysis studies using a simple modified blood film technique.

Water movement across the cell membrane is crucial, with red blood cells (RBCs) experiencing the flow of water in both directions at a rate of approximately 100 times their volume per second. This process typically results in no net water flow due to an equal balance of water movement in opposite directions, a phenomenon known as osmosis, driven by water potential or impermeant solute concentration. Understanding osmosis is essential for both physiology and medical practice, yet its complexity may not be effectively conveyed to the students through traditional teaching methods. This study presents a novel approach to observing the osmotic effect on RBCs using a simple, modified blood film technique. Aimed at enhancing educational understanding of cellular behavior in different osmotic environments, this method provides a practical hands-on learning experience. By applying various osmotic solutions to prepared blood films and observing the resultant morphological changes in RBCs under a microscope, this technique allows for direct visualization of osmosis in action.NEW & NOTEWORTHY This study presents an innovative teaching approach for understanding osmosis and its effects on red blood cells. Using a simple, modified blood film technique, students can visually observe and engage with the dynamic process of osmosis. This hands-on method enhances learning, making complex physiological concepts accessible and practical. Ideal for resource-limited settings, it bridges theoretical knowledge and practical application, transforming physiology education.

Osmotic Pressure and Its Biological Implications.

Gaining insight into osmotic pressure and its biological implications is pivotal for revealing mechanisms underlying numerous fundamental biological processes across scales and will contribute to the biomedical and pharmaceutical fields. This review aims to provide an overview of the current understanding, focusing on two central issues: (i) how to determine theoretically osmotic pressure and (ii) how osmotic pressure affects important biological activities. More specifically, we discuss the representative theoretical equations and models for different solutions, emphasizing their applicability and limitations, and summarize the effect of osmotic pressure on lipid phase separation, cell division, and differentiation, focusing on the mechanisms underlying the osmotic pressure dependence of these biological processes. We highlight that new theory of osmotic pressure applicable for all experimentally feasible temperatures and solute concentrations needs to be developed, and further studies regarding the role of osmotic pressure in other biological processes should also be carried out to improve our comprehensive and in-depth understanding. Moreover, we point out the importance and challenges of developing techniques for the in vivo measurement of osmotic pressure.

Transcriptional responses to salinity-induced changes in cell wall morphology of the euryhaline diatom Pleurosira laevis.

Diatoms are unicellular algae with morphologically diverse silica cell walls, which are called frustules. The mechanism of frustule morphogenesis has attracted attention in biology and nanomaterials engineering. However, the genetic regulation of the morphology remains unclear. We therefore used transcriptome sequencing to search for genes involved in frustule morphology in the centric diatom Pleurosira laevis, which exhibits morphological plasticity between flat and domed valve faces in salinity 2 and 7, respectively. We observed differential expression of transposable elements (TEs) and transporters, likely due to osmotic response. Up-regulation of mechanosensitive ion channels and down-regulation of Ca2+-ATPases in cells with flat valves suggested that cytosolic Ca2+ levels were changed between the morphologies. Calcium signaling could be a mechanism for detecting osmotic pressure changes and triggering morphological shifts. We also observed an up-regulation of ARPC1 and annexin, involved in the regulation of actin filament dynamics known to affect frustule morphology, as well as the up-regulation of genes encoding frustule-related proteins such as BacSETs and frustulin. Taken together, we propose a model in which salinity-induced morphogenetic changes are driven by upstream responses, such as the regulation of cytosolic Ca2+ levels, and downstream responses, such as Ca2+-dependent regulation of actin dynamics and frustule-related proteins. This study highlights the sensitivity of euryhaline diatoms to environmental salinity and the role of active cellular processes in controlling gross valve morphology under different osmotic pressures.

A quadriphasic mechanical model of the human dermis.

The present study investigates the multiphasic nature of the mechanical behavior of human dermis. Motivated by experimental observations and by consideration of its composition, a quadriphasic model of the dermis is proposed, distinguishing solid matrix components, interstitial fluid and charged constituents moving within the fluid, i.e., anions and cations. Compression and tensile experiments with and without change of osmolarity of the bath are performed to characterize the chemo-mechanical coupling in the dermis. Model parameters are determined through inverse analysis. The computations predict a dominant role of the permeability in the determination of the temporal evolution of the mechanical response of the tissue. In line with the previous studies on other tissues, the analysis shows that an ideal model based on Donnan's equilibrium overestimates the osmotic pressure in skin for the case of very dilute solutions. The quadriphasic model is applied to predict changes in dermal cell environment and therefore alterations in what is called the "mechanome," associated with skin stretch. The simulations indicate that skin deformation causes a variation in several local variables, including in particular the electric field associated with a deformation-induced non-homogeneous distribution of fixed charges.

Membrane behavior of clay under mixed solution conditions.

Compacted clay is employed as the buffer material for landfills, and multiple ions are dissolved in the leachate restricted by the compacted clay layer. The membrane efficiency is an important indicator to assess the barrier properties of the compacted clay layer and is measured through membrane tests. However, most membrane tests are currently conducted with a single solute solution, which does not reflect the mixed solution characteristics of leachates. To assess the membrane efficiency of compacted clay under mixed solution conditions, 13 membrane tests were conducted on a bentonite-amended soil using KCl-NaCl mixed solutions, KCl-CaCl2 mixed solutions, and KCl-AlCl3 mixed solutions with different mixing ratios at a total concentration of 20 mM. Nuclear magnetic resonance (NMR) tests were conducted on the soil specimen after the membrane tests to investigate the micromechanism of the membrane behavior under mixed solution conditions. Results indicate that the membrane efficiency increased with the mixing ratio of Na+ but decreased with the mixing ratio of Ca2+ or Al3+. In the 13 membrane tests, the lowest membrane efficiency was achieved when the specimen was tested with pure AlCl3 solution. The relationship between the membrane efficiency and mixing ratio was also investigated at the microscopic scale. As the ion valence increases, the diffuse double layer thickness is smaller and the proportion of macropores is larger, as verified by NMR tests.

Optimization of Zwitterionic Polymers for Cell Cryopreservation.

Cryopreservation techniques are valuable for the preservation of genetic properties in cells, and the development of this technology contributes to various fields. In a previous study, an isotonic freezing medium composed of poly(zwitterion) (polyZI) has been reported, which alleviates osmotic shock, unlike typical hypertonic freezing media. In this study, the primitive freezing medium composed of emerging polyZI is optimized. Imidazolium/carboxylate-type polyZI (VimC3 C) is the optimal chemical structure. The molecular weight and degree of ion substitution (DSion ) are not significant factors. There is an impediment with the primitive polyZI freezing media. While the polyZI forms a matrix around the cell membrane to protect cells, the matrix is difficult to remove after thawing, resulting in low cell proliferation. Unexpectedly, increasing the poly(VimC3 C) concentration from 10% to 20% (w/v) improves cell proliferation. The optimized freezing medium, 20% (w/v) poly(VimC3 C)_DSion(100%) /1% (w/v) NaCl aqueous solution, exhibited a better cryoprotective effect.

Osmotic stress induces the formation of migrasome-like vesicles.

Migrasomes are extracellular vesicles that form on the retraction fibers of migrating cells. In this study, we report the formation of migrasome-like vesicles enriched in tetraspanin 4 and containing cytoplasmic components in response to hypoosmotic stress. When migrating cells were subjected to hypoosmotic stress, vesicles with a size distribution of 0.5 to 2 µm formed on the retraction fibers, and vanished in a few minutes. The vesicles are rich in cholesterol, and their number was reduced when cells were pretreated with lipoprotein-deficient serum. The formation of migrasome-like vesicles upon hypoosmotic stress may provide biophysical cues regarding the cellular response to this external stimulus in cells and tissues.

The Changes in Relative Blood Volume during Dialysis Are Characterized by Ultrafiltration Rate and Predialysis Blood Test Values.

INTRODUCTION: Continuous monitoring of relative blood volume (percentage BV) in hemodialysis (HD) is critical for determining dry weight and preventing intradialytic hypotension. However, the cause of the BV variation remains unknown. This research aimed to examine factors that influence the percentage BV. METHODS: We devised a formula based on coefficients ("a," "τ," and "b") to predict changes in percentage BV. "a" denotes a significant decrease in percentage BV in the early stages of HD. "τ" represents the transition from early to late phase of HD. "b" denotes the slope of the decrease in percentage BV in the late phase of HD. We measured the percentage BV in 18 patients with end-stage renal disease. The coefficients were estimated by fitting experimental data from patients using a least squares optimization algorithm. A correlation analysis of these parameters and patient predialysis data was performed. RESULTS: Ultrafiltration rate (UFR) was found to be negatively correlated with "b" (r = -0.851, p < 0.01). However, UFR was not significantly related to "a." Predialysis serum total protein level was negatively correlated with "a" (r = -0.531, p = 0.042). Predialysis serum albumin and predialysis sodium were not significantly correlated with "a" and "τ." Plasma osmolarity did not have a significant relationship with "a" and "τ." DISCUSSION/CONCLUSION: UFR influenced the decrease in percentage BV in the late phase but did not influence the decrease of percentage BV in the early phase. "a" was associated with predialysis serum total protein level but not with plasma osmolality or predialysis sodium. This implies that colloid oncotic pressure is important for plasma refilling immediately after dialysis begins. During the change of percentage BV, the decrease in the early phase of dialysis was not related to UFR, but related to other parameters, especially predialysis total protein level. A decrease in the late phase of dialysis is related to UFR.

Effect of membrane tension on antimicrobial peptide PGLa-induced pore formation in lipid bilayers.

The osmotic pressure (Π) method has recently been developed to quantitatively examine the effect of membrane tension (σ) on pore formation in giant unilamellar vesicles (GUVs) induced by antimicrobial peptides (AMPs). Here, we used the Π method to reveal the effect of σ on the interaction of an AMP, PGLa, with lipid bilayers comprising dioleoylphosphatidylglycerol (DOPG) and dioleoylphosphatidylcholine (DOPC) (4/6). PGLa induced leakage of fluorescent probes from single GUVs under Π, indicating nanopore formation. Membrane tension did not transform a PGLa-induced nanopore into a micropore nor cause GUV burst up to 3.4 mN/m, which is in contrast with the effect of σ on another AMP, magainin 2-induced pore formation, where lower σ resulted in GUV burst. The fraction of leaking GUVs at a specific time increased with increasing σ, indicating that the rate of PGLa-induced pore formation increases with increasing σ. The rate of transfer of fluorescent probe-labeled PGLa across the lipid bilayer without pore formation also increased with increasing σ. PGLa-induced pore formation requires a symmetric distribution of peptides in both leaflets of the GUV bilayer, and thus we infer that the increase in the rate of PGLa transfer from the outer leaflet to the inner leaflet underlies the increase in the rate of pore formation with increasing σ. On the basis of these results, we discuss the difference between the effect of σ on nanopore formation in GUV membranes induced by PGLa and that by magainin 2.

Synergistic effects of microplastic and lead trigger physiological and biochemical impairment in a mangrove crab.

Microplastics (MP) are vectors for other environmental contaminants, such as metals, being a considerable problem, especially in the aquatic ecosystem. To investigate the combined effects of MP (high density polyethylene) with lead (Pb), we exposed the mangrove fiddler crab Minuca vocator to Pb (50 mg L-1), and MP (25 mg L-1) alone and in mixture, for 5 days. We aimed to determine Pb and MP bioaccumulation, as well as physiological (oxygen consumption and hemolymph osmolality) and biochemical (superoxide dismutase, catalase, glutathione peroxidase, and lipid peroxidation) traits effects. Co-exposure of MP and Pb significantly increased the bioaccumulation of Pb, but reduced MP tissue accumulation. Regarding the physiological traits, increasing osmolality and oxygen consumption rates compared to the control were observed, particularly in the combined Pb and MP exposure. As to biochemical traits, the combination of Pb and MP induced the most significant responses in the enzymatic profile antioxidant enzyme activity. The catalase (CAT), glutathione peroxidase (GPx), and dismutase superoxide (SOD) decreased compared to individual exposure effects; the combination of MP and Pb had a synergistic effect on promoting lipid peroxidation (LPO). The co-exposure of MP and Pb acted synergistically when compared to the effects of the isolated compounds. Due to the increasing MP contamination in mangroves, more severe physiological and biochemical effects can be expected on mangrove crabs exposed to metal contamination.

Characteristics of rice starches modified by single and dual heat moisture and osmotic pressure treatments.

The effect of osmotic pressure treatment (OPT), heat moisture treatment (HMT), and their dual combination as HMT-OPT and OPT-HMT on functional and pasting properties, gel texture, crystallinity, thermal, morphological, and rheological properties, and in vitro digestibility of modified starches were investigated. HMT was done with 29 % moisture at 111 °C for 45 min while OPT was performed at 117 °C for 35 min with saturated sodium sulphate solution. All modifications increased amylose content, improved pasting stability, and reduced swelling power and solubility. Dual modifications caused higher morphological changes than single modified starches. HMT and OPT increased pasting temperature, setback and final viscosity while decreased peak viscosity and breakdown, whereas HMT-OPT and OPT-HMT reduced all pasting parameters except pasting temperature. 1047/1022 and 995/1022 ratios and relative crystallinity decreased. V-type polymorphs were formed, and gelatinization temperature range increased with lower gelatinization enthalpy. Starch gel elasticity, RS and SDS content were enhanced to a greater extent after HMT-OPT and OPT-HMT. HMT as a single and dual form with OPT showed prominent effect on pasting, thermal, crystalline, and rheological properties. Application of HMT, OPT and dual modified starches with improved functionalities may be targeted for suitable food applications such as noodles.

Metabolites, ions, and the mechanisms behind seasonal cold hardening of Pyrochroa coccinea (Pyrochroidae) larvae.

The larvae of the black headed cardinal beetle Pyrochroa coccinea, overwinters under the bark of dead logs in northern European dioecious forests, and are thus exposed to temperatures below the melting point of their bodily fluids. Here we explore the mechanisms behind their seasonal cold hardening by characterising field samples collected monthly throughout the year. Both the lower lethal temperature and supercooling point dropped as much as 10℃ in the second half of November, reaching values around -15℃ by the beginning of December. This change was accompanied by a 320 mosmol/kg increase in hemolymph osmolality, which is a doubling compared to the summer levels. We used NMR metabolomics to identify and measure the absolute concentrations of the responsible cryoprotective C-H containing metabolites in the hemolymph. The largest increase was found to be in either glucose or trehalose, with an average total increase of 120 mM. Proline, alanine, and choline concentrations were found to increase by around 10 mM each. Contrarily, phosphocholine and phosphoethanolamine were halved, resulting in a total decrease of around 50 mM. These measurements were complemented with ion exchange chromatography measurements. This allowed us to account for all the osmotic pressure in the summer hemolymph, and the measured concentration changes explained as much as 40 % of the observed osmolality increase upon cold hardening. Preliminary results indicate that the remainder may be explained by non-colligative protein contributions.

Achilles' Heel-The Significance of Maintaining Microenvironmental Homeostasis in the Nucleus Pulposus for Intervertebral Discs.

The dysregulation of intracellular and extracellular environments as well as the aberrant expression of ion channels on the cell membrane are intricately linked to a diverse array of degenerative disorders, including intervertebral disc degeneration. This condition is a significant contributor to low back pain, which poses a substantial burden on both personal quality of life and societal economics. Changes in the number and function of ion channels can disrupt the water and ion balance both inside and outside cells, thereby impacting the physiological functions of tissues and organs. Therefore, maintaining ion homeostasis and stable expression of ion channels within the cellular microenvironment may prove beneficial in the treatment of disc degeneration. Aquaporin (AQP), calcium ion channels, and acid-sensitive ion channels (ASIC) play crucial roles in regulating water, calcium ions, and hydrogen ions levels. These channels have significant effects on physiological and pathological processes such as cellular aging, inflammatory response, stromal decomposition, endoplasmic reticulum stress, and accumulation of cell metabolites. Additionally, Piezo 1, transient receptor potential vanilloid type 4 (TRPV4), tension response enhancer binding protein (TonEBP), potassium ions, zinc ions, and tungsten all play a role in the process of intervertebral disc degeneration. This review endeavors to elucidate alterations in the microenvironment of the nucleus pulposus during intervertebral disc degeneration (IVDD), with a view to offer novel insights and approaches for exploring therapeutic interventions against disc degeneration.

Effect of Diacylglycerol Crystallization on W/O/W Emulsion Stability, Controlled Release Properties and In Vitro Digestibility.

Water-in-oil-in-water (W/O/W) emulsions with high-melting diacylglycerol (DAG) crystals incorporated in the oil droplets were fabricated and the compositions were optimized to achieve the best physical stability. The stability against osmotic pressure, encapsulation efficiency and in vitro release profiles of both water- and oil-soluble bioactives were investigated. The presence of interfacial crystallized DAG shells increased the emulsion stability by reducing the swelling and shrinkage of emulsions against osmotic pressure and heating treatment. DAG crystals located at the inner water/oil (W1/O) interface and the gelation of the inner phase by gelatin helped reduce the oil droplet size and slow down the salt release rate. The DAG and gelatin-contained double emulsion showed improved encapsulation efficiency of bioactives, especially for the epigallocatechin gallate (EGCG) during storage. The double emulsions with DAG had a lower digestion rate but higher bioaccessibility of EGCG and curcumin after in vitro digestion. DAG-stabilized double emulsions with a gelled inner phase thus can be applied as controlled delivery systems for bioactives by forming robust interfacial crystalline shells.