PLA plastic particles disrupt bile acid metabolism leading to hepatic inflammatory injury in male mice.

Microplastics, such as polylactic acid (PLA), are ubiquitous environmental pollutants with unclear implications for health impact. This study aims to elucidate the mechanisms of PLA-induced inflammatory liver injury, focusing on disturbance of bile acid metabolism. The in vitro PLA exposure experiment was conducted using HepG2 cells to assess cell viability, cytokine secretion, and effects on bile acid metabolism. In vivo, male C57BL/6 J mice were exposed to PLA for ten days continuously, liver function and histopathological assessment were evaluated after the mice sacrificed. Molecular analyses including quantitative reverse transcription polymerase chain reaction (RT-qPCR) and Western blotting, were applied to evaluate the expression of bile acid metabolizing enzymes and transporters. PLA exposure resulted in decreased cell viability in HepG2 cells, increased inflammation and altered bile acid metabolism. In mice, PLA exposure resulted in decreased body weight and food intake, impaired liver function, increased hepatic inflammation, altered bile acid profiles, and dysregulated expression of bile acid metabolic pathways. PLA exposure disrupts bile acid metabolism through inhibition of the CYP7A1 enzyme and activation of the FGF-JNK/ERK signaling pathway, contributing to liver injury. These findings highlight the potential hepatotoxic effects of environmentally friendly plastics PLA and underscore the need for further research on their biological impact.

Contrast Enhanced Ultrasonography of Kidney in Chronic Intermittent Hypoxia Rat Model.

OBJECTIVES: The objective of our study was to assess the ability of contrast-enhanced ultrasound (CEUS) in evaluating renal microperfusion in an animal model. METHODS: Twenty Sprague-Dawley rats were subdivided into two groups: the normal and chronic intermittent hypoxia (CIH) groups. In the CIH model, 10 Sprague-Dawley rats were exposed to CIH for 8 weeks to mimic obstructive sleep apnea (OSA). The CEUS parameters of the renal cortex and medulla were obtained and compared between groups. The pathological changes of the kidney tissues were examined by histological staining such as hematoxylin and eosin (H&E) and Masson's trichrome. RESULTS: CIH caused morphological damage to kidneys. In the cortex, the peak intensity (PI) (P = .009) was significantly lower and time to peak (Ttop) (P = .019) was significantly prolonged in the CIH group compared with the controls. The area under ascending curve (WiAUC) in the medulla and cortex were both significantly lower in the CIH group than those in the control group (P both <.05). CEUS parameters (including PI and WiAUC of the cortex and WiAUC of the medulla) were negatively correlated with serum creatinine (P all <.05). In the medulla, the area under descending curve (WoAUC) was positively correlated with serum creatinine (P = .027), PI was negatively correlated with uric acid (P = .034). CONCLUSION: CEUS parameters (including Ttop, PI, WoAUC, and WiAUC) reflect renal microvascular changes in CIH. CEUS could be a safe and accurate imaging method to assess renal microvascular damage in CIH rats.

Dlg1 Maintains Dendritic Cell Function by Securing Voltage-Gated K+ Channel Integrity.

Dendritic cells (DCs) play key roles in Ab responses by presenting Ags to lymphocytes and by producing proinflammatory cytokines. In this study, we reported that DC-specific knockout of discs large homologue 1 (Dlg1) resulted in a significantly reduced capacity to mediate Ab responses to both thymus-independent and thymus-dependent Ags in Dlg1 fl/flCd11c-Cre-GFP mice. Mechanistically, Dlg1-deficient DCs showed severely impaired endocytosis and phagocytosis capacities upon Ag exposure. In parallel, loss of Dlg1 significantly jeopardized the proinflammatory cytokine production by DCs upon TLR stimulation. Thus, Dlg1-deficient DCs lost their functions to support innate and adaptive immunities. At a cellular level, Dlg1 exhibited an indispensable function to maintain membrane potential changes by securing potassium ion (K+) efflux and subsequent calcium ion (Ca2+) influx events in DCs upon stimulation, both of which are known to be required for proper function of DCs. At a molecular level, Dlg1 did so by retaining the integrity of voltage-gated K+ channels (including Kv1.3) in DCs. The loss of Dlg1 led to a decreased expression of K+ channels, resulting in impaired membrane potential changes and, as a consequence, reduced proinflammatory cytokine production, compromised Ag endocytosis, and phagocytosis. In conclusion, this study provided, to our knowledge, a novel insight into Dlg1 and the voltage-gated K+ channels axis in DC functions.

Impaired maturation of large dense-core vesicles in muted-deficient adrenal chromaffin cells.

The large dense-core vesicle (LDCV), a type of lysosome-related organelle, is involved in the secretion of hormones and neuropeptides in specialized secretory cells. The granin family is a driving force in LDCV biogenesis, but the machinery for granin sorting to this biogenesis pathway is largely unknown. The mu mutant mouse, which carries a spontaneous null mutation on the Muted gene (also known as Bloc1s5), which encodes a subunit of the biogenesis of lysosome-related organelles complex-1 (BLOC-1), is a mouse model of Hermansky-Pudlak syndrome. Here, we found that LDCVs were enlarged in mu adrenal chromaffin cells. Chromogranin A (CgA, also known as CHGA) was increased in mu adrenals and muted-knockdown cells. The increased CgA in mu mice was likely due a failure to export this molecule out of immature LDCVs, which impairs LDCV maturation and docking. In mu chromaffin cells, the size of readily releasable pool and the vesicle release frequency were reduced. Our studies suggest that the muted protein is involved in the selective export of CgA during the biogenesis of LDCVs.

Advancements in mitochondrial-targeted nanotherapeutics: overcoming biological obstacles and optimizing drug delivery.

In recent decades, nanotechnology has significantly advanced drug delivery systems, particularly in targeting subcellular organelles, thus opening new avenues for disease treatment. Mitochondria, critical for cellular energy and health, when dysfunctional, contribute to cancer, neurodegenerative diseases, and metabolic disorders. This has propelled the development of nanomedicines aimed at precise mitochondrial targeting to modulate their function, marking a research hotspot. This review delves into the recent advancements in mitochondrial-targeted nanotherapeutics, with a comprehensive focus on targeting strategies, nanocarrier designs, and their therapeutic applications. It emphasizes nanotechnology's role in enhancing drug delivery by overcoming biological barriers and optimizing drug design for specific mitochondrial targeting. Strategies exploiting mitochondrial membrane potential differences and specific targeting ligands improve the delivery and mitochondrial accumulation of nanomedicines. The use of diverse nanocarriers, including liposomes, polymer nanoparticles, and inorganic nanoparticles, tailored for effective mitochondrial targeting, shows promise in anti-tumor and neurodegenerative treatments. The review addresses the challenges and future directions in mitochondrial targeting nanotherapy, highlighting the need for precision, reduced toxicity, and clinical validation. Mitochondrial targeting nanotherapy stands at the forefront of therapeutic strategies, offering innovative treatment perspectives. Ongoing innovation and research are crucial for developing more precise and effective treatment modalities.

The Impact of NaOH on the Micro-Mechanical Properties of the Interface Transition Zone in Low-Carbon Concrete.

To tackle carbon emissions from cement production and address the decline in concrete's mechanical properties due to the substitution of cement with solid waste (glass powder) and natural mineral admixture (zeolite powder) materials, we employed glass powder and zeolite powder to create composite cementitious materials. These materials underwent alkali activation treatment with a 4% NaOH dosage, replacing 50% of cement to produce low-carbon concrete. Nanoindentation tests and mercury intrusion porosimetry (MIP) were employed to uncover the micro-mechanical properties and influencing mechanisms of alkali-activated low-carbon concrete. The results indicate a notable enhancement in the indentation modulus (19.9%) and hardness (25.9%) of alkali-activated low-carbon concrete compared to non-activated concrete. Simultaneously, the interfacial transition zone thickness decreased by 10 µm. The addition of NaOH led to a reduced volume fraction of pores (diameter >100 nm) and an increased fraction of pores (diameter < 100 nm), thereby reducing porosity by 2.6%, optimizing the pore structure of low-carbon concrete. The indentation modulus, hardness and volume fraction of the hydrated phase derived from Gaussian fitting analysis of the nanoindentation statistics showed that NaOH significantly improved the modulus and hardness of the hydration products of low-carbon concrete. This activation resulted in decreased LDC-S-H gel (low-density hydrated calcium silicate Ca5Si6O16(OH)·4H2O) and pore content, while the HD C-S-H gel (high-density hydrated calcium silicate Ca5Si6O16(OH)·4H2O) and CH (calcium hydroxide crystals Ca(OH)2) content increased by 13.91% and 23.46%, respectively. Consequently, NaOH influenced the micro-mechanical properties of low-carbon concrete by generating more high-density hydration products, reducing pore content, enhancing the pore indentation modulus and hardness, and shortening the interfacial transition zone. This study offers novel insights into reducing carbon emissions and promoting the use of solid waste (glass powder) and natural mineral admixture (zeolite powder) materials in concrete, contributing to the advancement of sustainable construction practices.

Mechanical Properties of Aeolian Sand Concrete Made from Alkali-Treated Aeolian Sand and Zeolite Powder.

The aim of this study is to promote the application of the excited zeolite powder (ZP)with aeolian sand powder (ASP) in the field of aeolian-sand concrete (ASC) production. This study utilises NaOH to treat composite cementitious materials containing aeolian sand and zeolite powders, which were used to replace 50% of the cement in aeolian-sand concrete (ASC). Production of alkali-inspired cement-based windswept concrete(AAZC).The mechanical properties of treated ASC considerably improved, especially when the NaOH dosage was 4% by mass. After curing this sample (denoted as AAZC-4) for 28 d, its compressive strength improved by 17.2%, and its split tensile increased by 16.3%. Potassium feldspar and montmorillonite in zeolite powder and SiO2 in the sand were decomposed by OH- and combined with other elements to generate various silicate gels and A-type potassium zeolite crystals inside the concrete. Microscopic examination showed that the gels and crystals intertwined to fill the pores, decreasing (increasing) the percentage of large (small) pores, thus optimising the pore structure. This substantially improved the mechanical properties of ASC. Freeze-thaw salt-intrusion tests showed that the extent of mass loss, degree of damage and loss of compressive strength of AAZC-4 were similar to those of ordinary concrete but were reduced by 36.8%, 19% and 52.1%, respectively, compared with those of ASC. Therefore, AAZC-4 has a sustainable working performance in chloride-ion permeable environments in cold and arid areas.

Lactoferrin alleviates chronic low­grade inflammation response in obese mice by regulating intestinal flora.

Chronic low­grade inflammation defines obesity as a metabolic disorder. Alterations in the structure of gut flora are strongly associated with obesity. Lactoferrin (LF) has a biological function in regulating intestinal flora. The present study aimed to investigate the therapeutic and anti­-inflammatory effects of LF in obese mice based on intestinal flora. A total of 30 C57BL/6 mice were divided into three groups consisting of 10 mice each. Subsequently, one group was fed a normal diet (Group K), another group was fed a high­fat diet (Group M) and the remaining group switched from regular drinking to drinking 2% LF water (Group Z2) after 2 weeks of high­fat diet; all mice were fed for 12 weeks. After the experiment, the mouse blood lipid and lipopolysaccharide levels, levels of inflammatory factors and intestinal tight junction proteins were assessed. Mouse stool samples were analyzed using 16S ribosomal RNA sequencing. The results showed that LF reduced serum total cholesterol, triglycerides and low­density lipoprotein levels, elevated high­density lipoprotein levels, suppressed metabolic endotoxemia and attenuated chronic low­grade inflammatory responses in obese mice. In addition, LF upregulated zonula occludens­1 and occludin protein expression levels in the intestine, thereby improving intestinal barrier integrity. LF altered the intestinal microbial structure of obese mice, reduced the ratio of Firmicutes and an elevated ratio of Bacteroidota, modifying the bacterial population to the increased relative abundance of Alistipes, Acidobacteriota, Psychrobacter and Bryobacter.

RNA Interference Reveals the Impacts of CYP6CY7 on Imidacloprid Resistance in Aphis glycines.

Cytochrome P450 (CYP) is a group of important detoxification enzymes found in insects related to their resistance to insecticides. To elucidate the CYP6 family genes of P450, which are potentially related to imidacloprid resistance in Aphis glycines, the CYP6 cDNA sequences of A. glycines were studied. The transcriptome of A. glycines was constructed, and the CYP6 cDNA sequences of A. glycines were screened. Their relative expression levels in response to imidacloprid induction were examined through qRT-PCR, and the CYP6s with higher expression levels were used to study the detoxification of imidacloprid through RNA interference and a bioassay. Twelve CYP6s were obtained from the A. glycines transcriptome. These samples were named by the International P450 Nomenclature Committee and registered in GenBank. After 3, 6, 12, 24 and 48 h of induction with LC50 concentrations of imidacloprid, the relative expression levels of these CYP6s increased; the expression level of CYP6CY7 experienced the highest increase, being more than 3-fold higher than that of those of the non-imidacloprid-induced CYP6s. After RNA interference for CYP6CY7, the relative expression level of CYP6CY7 significantly decreased after 3, 6 and 12 h, while the corresponding P450 enzyme activity decreased after 12 and 24 h. The mortality of A. glycines due to imidacloprid treatment increased by 14.71% at 24 h. CYP6CY7 might detoxify imidacloprid in A. glycines. This study provides a theoretical basis for the further study of the mechanism of action of CYP6s and potential new methods for improving insecticidal efficacy.

Investigating the modulatory effects of lactoferrin on depressed rats through 16S rDNA gene sequencing and LC-MS metabolomics analysis.

Lactoferrin is a natural multifunctional glycoprotein with potential antidepressant-like effects. However, the mechanism of its antidepressant effect has not been explored from the perspective of gut flora metabolism. Therefore, we employed both 16S rDNA gene sequencing and LC-MS metabolomics analysis to investigate the regulatory effects and mechanisms of lactoferrin in a rat model of depression. After one week of acclimatization, twenty-four 7-week-old male Sprague-Dawley rats were randomly and equally assigned into three groups: the control group, the model group, and the lactoferrin intervention group. The control group rats were housed under standard conditions, while the rats in the model and lactoferrin intervention groups were individually housed and exposed to chronic unpredictable mild stress for 44 days simultaneously. The lactoferrin intervention group was provided with water containing 2% lactoferrin (2 g/100 ml). Behavioural tests were conducted at week 7. Upon completion of the behavioral tests, the rats were anesthetized with isoflurane, humanely euthanized using a rat guillotine, and tissue samples were collected for further experiments. The results indicated that lactoferrin intervention led to an increase in sucrose solution consumption, horizontal movement distance, number of cross platforms, and residence time in the target quadrant. Additionally, it resulted in an increase in jejunal tight junction protein ZO-1 expression and a suppression of serum expression of inflammatory factors, Lipopolysaccharide and Diamine oxidase. In summary, lactoferrin can regulate the metabolic disorder of intestinal flora, reduce intestinal permeability, and further regulate the metabolic balance of hippocampal tissues through the microbiota-gut-brain axis. This process ultimately alleviates the depression-like behavior in rats.

Clathrin mediates membrane fission and budding by constricting membrane pores.

Membrane budding, which underlies fundamental processes like endocytosis, intracellular trafficking, and viral infection, is thought to involve membrane coat-forming proteins, including the most observed clathrin, to form Ω-shape profiles and helix-forming proteins like dynamin to constrict Ω-profiles' pores and thus mediate fission. Challenging this fundamental concept, we report that polymerized clathrin is required for Ω-profiles' pore closure and that clathrin around Ω-profiles' base/pore region mediates pore constriction/closure in neuroendocrine chromaffin cells. Mathematical modeling suggests that clathrin polymerization at Ω-profiles' base/pore region generates forces from its intrinsically curved shape to constrict/close the pore. This new fission function may exert broader impacts than clathrin's well-known coat-forming function during clathrin (coat)-dependent endocytosis, because it underlies not only clathrin (coat)-dependent endocytosis, but also diverse endocytic modes, including ultrafast, fast, slow, bulk, and overshoot endocytosis previously considered clathrin (coat)-independent in chromaffin cells. It mediates kiss-and-run fusion (fusion pore closure) previously considered bona fide clathrin-independent, and limits the vesicular content release rate. Furthermore, analogous to results in chromaffin cells, we found that clathrin is essential for fast and slow endocytosis at hippocampal synapses where clathrin was previously considered dispensable, suggesting clathrin in mediating synaptic vesicle endocytosis and fission. These results suggest that clathrin and likely other intrinsically curved coat proteins are a new class of fission proteins underlying vesicle budding and fusion. The half-a-century concept and studies that attribute vesicle-coat contents' function to Ω-profile formation and classify budding as coat-protein (e.g., clathrin)-dependent or -independent may need to be re-defined and re-examined by considering clathrin's pivotal role in pore constriction/closure.

How micron-sized exocrine vesicles release content: A comparison with sub-micron endocrine vesicles.

Exocytosis releases vesicular contents to mediate physiological functions. In this issue, Biton et al. (https://doi.org/10.1083/jcb.202302112) found four modes of releasing micron-sized exocrine vesicles and the underlying mechanisms involving actomyosin and BAR domain proteins. We highlight their discovery, compare it with much smaller/faster neuroendocrine vesicle fusion, and draw distinct and conserved principles regarding their membrane transformations, pore dynamics, and underlying mechanisms.

Highly Breathable, Stretchable, and Tailorable TPU Foam for Flexible Gas Sensors.

Continuous real-time monitoring of air quality is of great significance in the realms of environmental monitoring, personal safety, and healthcare. Recently, flexible gas sensors have gained great popularity for their potential to be integrated into various smart wearable electronics and display devices. However, the development of gas sensors with superior sensitivity, breathability, and stretchability remains a challenge. Here, a new high porosity thermoplastic polyurethane (HP-TPU) foam was reported for gas sensors, which exhibited large three-dimensional network structures and excellent mechanical properties. The HP-TPU foam was achieved by using a simple steam-induced method, which was suitable for mass production. The unique structure endowed this foam with 77.5% porosity, 260% strain ability, and 0.45 MPa Young's modulus, which improved 35, 31, and 80%, respectively, compared to previously reported traditional TPU foam (T-TPU) prepared by the drying method. In addition, the foam presented high gas permeability (312 g/m-2, 24 h) and excellent stability, and it remained undamaged even after 2000 cycles at 70% strain. The sensing material was coated on a PET flexible interdigital electrode and sandwiched between two HP-TPU foam layers for a gas sensitivity test. Due to the easy diffusion of gas between the pores and contact with the sensing materials, the HP-TPU foam exhibited a significant reduction of 85% in average response time and 46% in average recovery time, compared to the T-TPU foam. A wearable sensing device, comprising sensing, data processing, and wireless transmission modules, was successfully developed to enable outdoor testing and achieved a detection range at the ppb level. Finally, the cytotoxicity test results confirmed that this flexible gas sensor did not harm human health. These results proved that this HP-TPU foam was an ideal matrix for the flexible gas sensor, exhibiting great application potential in the fields of seamless human-machine integration.

Chronic Unpredictable Mild Stress in Rats based on the Mongolian medicine.

Depression is a prevalent affective disorder and constitutes a leading cause of global disability. The limitations of current pharmacological interventions contribute to the substantial health burden attributed to this condition. There is a pressing need for a deeper understanding of the underlying mechanisms of depression, making pre-clinical models with translational potential highly valuable. Mongolian medicine, a subset of traditional medicine, posits that disease occurrence is closely tied to the equilibrium of wind, bile, and Phlegm. In this study, we introduce a protocol for the chronic unpredictable mild stress (CUMS) method in rats. Within this framework, rats are subjected to a series of fluctuating, mild stressors to induce a depression-like phenotype, mimicking the pathogenesis of human depression. Behavioral assays employed in this protocol include the sucrose preference test (SPT), indicative of anhedonia-a core symptom of depression; the open field test (OFT), which measures anxiety levels; and the Morris water maze test (MWM), which evaluates spatial memory and learning abilities. The CUMS method demonstrates the capability to induce anhedonia and to cause long-term behavioral deficits. Furthermore, this protocol is more aligned with Mongolian medical theory than other animal models designed to elicit depression-like behavior. The development of this animal model and subsequent research provide a robust foundation for future innovative studies in the realm of Mongolian medicine.

Real-time visualization of exo- and endocytosis membrane dynamics with confocal and super-resolution microscopy.

Real-time confocal and super-resolution imaging reveals membrane dynamics of exo- and endocytosis, including hemi-fusion, fusion pore opening, expansion, constriction, closure (kiss-and-run), fused-vesicle shrinking (shrink fusion), and flat membrane transition to vesicles via intermediate Λ- and Ω-shape structures. Here, we describe a protocol for imaging these membrane dynamics, including primary culture of bovine adrenal chromaffin cells, fluorescent probe application, patch-clamp to deliver depolarization and evoke exo- and endocytosis, electron microscopy (EM), and real-time confocal and stimulated emission depletion (STED) microscopy. For complete details on the use and execution of this protocol, please refer to Zhao et al. (2016), Shin et al. (2018), and Shin et al. (2021).

Sequential compound fusion and kiss-and-run mediate exo- and endocytosis in excitable cells.

Vesicle fusion at preestablished plasma membrane release sites releases transmitters and hormones to mediate fundamental functions like neuronal network activities and fight-or-flight responses. This half-a-century-old concept-fusion at well-established release sites in excitable cells-needs to be modified to include the sequential compound fusion reported here-vesicle fusion at previously fused Ω-shaped vesicular membrane. With superresolution STED microscopy in excitable neuroendocrine chromaffin cells, we real-time visualized sequential compound fusion pore openings and content releases in generating multivesicular and asynchronous release from single release sites, which enhances exocytosis strength and dynamic ranges in excitable cells. We also visualized subsequent compound fusion pore closure, a new mode of endocytosis termed compound kiss-and-run that enhances vesicle recycling capacity. These results suggest modifying current exo-endocytosis concepts by including rapid release-site assembly at fused vesicle membrane, where sequential compound fusion and kiss-and-run take place to enhance exo-endocytosis capacity and dynamic ranges.

[Controlled syntheses and photoluminescence characterization of monoclinic cerium orthophosphate with nanostructure].

The nanostructured CePO4 with monoclinic phase was controllably synthesized through a low temperature hydrothermal route by varying the reactant PO4(3-)/Ce(3+) molar ratio. The structures, morphologies, sizes and luminescence properties of the products were studied by XRD, FE-SEM, DSC-TG and photoluminescence spectra. With the PO4(3-)/Ce(3+) molar ratios increased, the synthesis temperature of as-synthesized monoclinic CePO4 was decreased, and the morphologies underwent the evolution from the rod-like nano-structures to the flower-like nanoclusters. When the PO4(3-)/Ce(3+) molar ratio was lower, CePO4 nanorods were obtained, while the PO4(3-)/Ce(3+) molar ratio was higher, the monoclinic CePO4 flower-like nanoclusters were crystallized. The photoluminescence spectrum showed that the CePO4 nanorods exhibit better photoluminescent property than the CePO4 flower-like nanoclusters. With the cycling use of phosphoric acid, the low-cost preparation of CePO4 could be achieved.

Hyperglycemia-Induced Dysregulated Fusion Intermediates in Insulin-Secreting Cells Visualized by Super-Resolution Microscopy.

Impaired insulin release is a hallmark of type 2 diabetes and is closely related to chronically elevated glucose concentrations, known as "glucotoxicity." However, the molecular mechanisms by which glucotoxicity impairs insulin secretion remain poorly understood. In addition to known kiss-and-run and kiss-and-stay fusion events in INS-1 cells, ultrafast Hessian structured illumination microscopy (Hessian SIM) enables full fusion to be categorized according to the newly identified structures, such as ring fusion (those with enlarged pores) or dot fusion (those without apparent pores). In addition, we identified four fusion intermediates during insulin exocytosis: initial pore opening, vesicle collapse, enlarged pore formation, and final pore dilation. Long-term incubation in supraphysiological doses of glucose reduced exocytosis in general and increased the occurrence of kiss-and-run events at the expense of reduced full fusion. In addition, hyperglycemia delayed pore opening, vesicle collapse, and enlarged pore formation in full fusion events. It also reduced the size of apparently enlarged pores, all of which contributed to the compromised insulin secretion. These phenotypes were mostly due to the hyperglycemia-induced reduction in syntaxin-1A (Stx-1A) and SNAP-25 protein, since they could be recapitulated by the knockdown of endogenous Stx-1A and SNAP-25. These findings suggest essential roles for the vesicle fusion type and intermediates in regulating insulin secretion from pancreatic beta cells in normal and disease conditions.

Physiological evidence of mitochondrial permeability transition pore opening caused by lipid deposition leading to hepatic steatosis in db/db mice.

Mitochondrial permeability transition pore (mPTP) is an important regulator in cell apoptosis and necrosis. However, its role in hepatic steatosis, especially its electrophysiological properties transformation remains elusive. Herein, using diabetes mice, we investigated the role of mPTP in hepatic steatosis triggered by diabetes and the mechanisms involved. We found that hepatic steatosis altered mitochondrial morphology, generating mega mitochondria, mitochondria swelling, calcein fluorescence quenching and mitochondrial membrane potential depolarization. At the same time, we confirmed an augmented mPTP opening with patch clamping in liver mitoplasts in db/db mice and a similar transformation with arachidonic acid (AA) simulating liquid deposition. We also found mPTP opening was significantly attenuated in wt mice after removing mitochondrial matrix, while that in db/db mice remained active. In addition, we observed that AA could directly activate mPTP in inside-out mode, independent of matrix calcium. In conclusion, we for the first time provided a physiological evidence of mPTP opening in lipid deposition, which could be directly induced by AA without Ca2+ and can be inhibited by cyclosporine A. As a result, it led to mitochondria morphology and function transformation. This might provide insights into potential therapeutic target for future treatment of mitochondrial liver disease.

Preformed Ω-profile closure and kiss-and-run mediate endocytosis and diverse endocytic modes in neuroendocrine chromaffin cells.

Transformation of flat membrane into round vesicles is generally thought to underlie endocytosis and produce speed-, amount-, and vesicle-size-specific endocytic modes. Visualizing depolarization-induced exocytic and endocytic membrane transformation in live neuroendocrine chromaffin cells, we found that flat membrane is transformed into Λ-shaped, Ω-shaped, and O-shaped vesicles via invagination, Λ-base constriction, and Ω-pore constriction, respectively. Surprisingly, endocytic vesicle formation is predominantly from not flat-membrane-to-round-vesicle transformation but calcium-triggered and dynamin-mediated closure of (1) Ω profiles formed before depolarization and (2) fusion pores (called kiss-and-run). Varying calcium influxes control the speed, number, and vesicle size of these pore closures, resulting in speed-specific slow (more than ∼6 s), fast (less than ∼6 s), or ultrafast (<0.6 s) endocytosis, amount-specific compensatory endocytosis (endocytosis = exocytosis) or overshoot endocytosis (endocytosis > exocytosis), and size-specific bulk endocytosis. These findings reveal major membrane transformation mechanisms underlying endocytosis, diverse endocytic modes, and exocytosis-endocytosis coupling, calling for correction of the half-a-century concept that the flat-to-round transformation predominantly mediates endocytosis after physiological stimulation.