American journal of physiology. Cell physiology最新文献

The aim of the present study was to investigate the secretory responses elicited by inositol 1,4,5-trisphosphate (IP₃) and their regulation by Ca²⁺ from different sources. Fura-2, carbon fiber amperometry, and plasma membrane capacitance recordings were performed in mouse chromaffin cells to evaluate cytosolic Ca²⁺ changes, catecholamine release, and exocytosis, respectively. Amperometric recordings revealed that IP₃ triggered the continuous release of catecholamines to the cytosol with a plateau shape, either applied independently or in combination with the V-ATPase blocker bafilomycin A1, without exhibiting additive effects, which suggests that V-ATPase blockade might be a potential mechanism of action. The catecholamine release elicited by IP₃ can take place in the absence of cytosolic Ca²⁺; however, it may be also regulated by it through a bell-shaped mechanism, with the contribution of Ca²⁺ stored in intracellular organelles. Furthermore, plasma membrane capacitance recordings showed that IP₃ could also elicit exocytosis of secretory vesicles with the participation of intracellular organelle Ca²⁺ stores. This exocytosis could be regulated by vesicular or cytosolic Ca²⁺, as shown in experiments with bafilomycin A1 or the Ca²⁺ chelator BAPTA-AM, respectively, and by kaempferol, an activator of the mitochondrial Ca²⁺ uniporter, suggesting that mitochondria may exert physiologically this Ca²⁺ regulatory mechanism. Therefore, in the IP₃-mediated secretion, Ca²⁺ from different sources control the different steps of catecholamine release from the secretory vesicle to the cytosol and then finally to the extracellular space.NEW & NOTEWORTHY Inositol 1,4,5-trisphosphate (IP₃) triggers the release of catecholamines from secretory vesicles to the cytosol through a process that may occur in the absence of cytosolic Ca²⁺, it is biphasically regulated by it and is dependent on Ca²⁺ from intracellular organelles. Additionally, IP₃ triggers the exocytosis of secretory vesicles through a cytosolic and vesicular Ca²⁺ regulatory mechanism that may be physiologically modulated by mitochondria.

Preeclampsia (PE) is a complex gestational disorder marked by vascular abnormalities and elevated blood pressure yet remains without widely effective treatments. This study investigates the efficacy of ferulic acid (FA) in alleviating PE symptoms by targeting the signal transducer and activator of transcription 3 (STAT3)/vascular endothelial growth factor (VEGF) signaling axis to enhance endothelial integrity and reduce inflammation. An N^G-nitro-l-arginine methyl ester hydrochloride (l-NAME)-induced PE mouse model was used, with FA administration to pregnant mice to assess therapeutic effects on key outcomes such as blood pressure, proteinuria, and placental function. Single-cell RNA sequencing (scRNA-seq) and molecular assays were conducted to examine FA's impact on endothelial cell balance, inflammation, and pathway-specific activity. The results showed that FA treatment significantly reduced hypertension, proteinuria, and inflammation, while improving endothelial cell balance in PE mice. In addition, inhibition of STAT3 phosphorylation by FA enhanced endothelial barrier function, stabilized vascular integrity, and supported improved fetal development outcomes. Overall, these findings demonstrate the protective effects of FA in PE by alleviating endothelial impairment and dampening inflammatory activity, offering a promising strategy to improve maternal and fetal health in PE, with implications for managing pregnancy-related vascular dysfunctions.NEW & NOTEWORTHY Our study investigates ferulic acid (FA) as a potential therapeutic intervention for preeclampsia (PE), a severe pregnancy complication with limited treatment options. By targeting the STAT3/VEGF signaling pathway, FA demonstrated significant reductions in hypertension, inflammation, and improved endothelial cell balance in PE mice. These results highlight FA's promise in enhancing maternal and fetal health by addressing endothelial dysfunction, suggesting its potential for broader applications in managing pregnancy-related vascular dysfunctions.

Amino acids (AAs) play structural and metabolic roles in muscle, heart, and liver-tissues impacted by cancer and chemotherapy. Changes in AA profiles within these tissues have not been evaluated in response to tumor growth and chemotherapy. This study investigated how tumor growth with or without doxorubicin altered tissue-level amino acids. Female C57bl/6 mice (n = 7-10/group) were randomly assigned to groups: control, doxorubicin control at 3 and 7 days, 21-day tumor, 24-day tumor, 28-day tumor, 24-day tumor + doxorubicin, 28-day tumor + doxorubicin. Tumor groups were injected with E0771 cells in the right flank on day 0. Doxorubicin was administered once (intraperitoneally) at 10 mg/kg in doxorubicin control and tumor + doxorubicin groups on day 21, with endpoints at day 24 and 28. Muscle glutamate and aspartate were significantly depleted by day 28 in both tumor and tumor + doxorubicin groups (P < 0.05), whereas proline, arginine, leucine, and isoleucine increased (P < 0.05). Hepatic aspartate was elevated by 21 days, and lysine by 24 days (P < 0.05). Cardiac glutamate was depleted at days 21, 24, and 28 (P < 0.05). Notably, doxorubicin did not add to tumor-induced changes in muscle or heart. Tumor AAs remained largely stable. Tumor growth induced profound changes to skeletal muscle AA pools, reflecting impaired handling of AAs that could serve structural roles, or expand the substrate pool for ATP synthesis. Despite this, most tumor AAs remained stable over tumor growth. These results suggest a link between muscle wasting and skeletal muscle-derived AAs for tumor growth. Further work is needed to characterize the mechanisms mediating the observed changes in AA profiles.NEW & NOTEWORTHY This study demonstrates significantly perturbed amino acid pools within muscle as a result of tumor growth, with marginal additive effects of doxorubicin administration. Notably, tumor amino acid pools remain primarily unchanged despite muscle suggesting significant changes, which may be indicative of structural damage or reduced ability to produce energy.

The transmembrane-electrostatically localized protons/cations charges (TELPs/TELCs) theory can serve as a theoretical framework to better explain cell electrophysiology and elucidate bioenergetic systems, including both delocalized and localized protonic coupling. According to the TELCs model, the excess positive charges of TELCs at one side of the membrane are balanced by the excess negative charges of transmembrane-electrostatically localized hydroxide anions (TELAs) at the other side of the membrane. Through the TELCs-membrane-TELAs capacitor model, the energetics of oxidative phosphorylation have recently been better elucidated in mitochondria and alkalophilic bacteria, leading to the identification of a novel Type-B energetic process. Both the TELCs model studies and experimental demonstration results showed that the putative "potential well/barrier" model is not needed to explain TELPs formation. Application of the TELCs model to neural cells has recently resulted in novel neural transmembrane potential integral equations. In this review article, we will visit the TELCs-membrane-TELAs model and its applications, including its features and predictions that may help better understand cell energetics. Meanwhile, we will also discuss some of the recent critiques and point out the opportunities and directions for future research. The TELCs model can be well predictive and provide new opportunities as a theoretical tool for further research to better understand cell physiology, bioenergetics, and neurosciences. This Landmark Review article timely provides the latest discoveries, breakthrough advances with new developments and knowledge, directions and opportunities for future research in a major emerging and exciting scientific area of protonic capacitor cell energetics: transmembrane-electrostatically localized protons/cations.

Eye disease-associated K⁺ channel Kir7.1 is highly expressed together with the Na⁺-K⁺ pump at the apical membrane of retinal pigment epithelial cells (RPEs) that line the subretinal space (SRS). SRS K⁺ concentration ([K⁺]_SRS) decreases from ∼5 to 2 mM upon light stimulation. Kir7.1 is crucial in its buffering, with failure thought to be causal in visual disease mutations of its gene. The unusual inverse relation to [K⁺]_o of its conductance, deemed essential for [K⁺]_SRS buffering, relies on nonconserved outer pore methionine-125. We now probe the role of Kir7.1 in the visual process by generating Kir7.1-M125R mutant mice with the channel predicted to lack [K⁺]_SRS buffering ability. RPE cell electrical properties and mouse electroretinograms (ERG) are assessed. Membrane potential of RPE cells was found to be dominated by K⁺, but while conductance decreased with increasing [K⁺]_o in control cells, the reverse was true for cells of Kir7.1-M125R-expressing mice. ERG of mutant animals revealed a larger c-wave than in controls, consistent with the relative K⁺ permeabilities of the RPE. In contrast, there was no difference between the a- and b-waves of Kir7.1-M125R and control mice, suggesting normal functioning of photoreceptors and bipolar cells, and therefore retinal processing of the light signal. If, as predicted, [K⁺]_SRS buffering is altered in mutant animals, this does not affect the retinal processing of the light signal. Other consequences of Kir7.1 malfunction, such as proposed function in photoreceptor outer segment recycling, must be involved in originating the disease phenotype associated with mutations in its gene.NEW & NOTEWORTHY Retinal pigment epithelium apical membrane K⁺ channel Kir7.1 is crucial in the buffering of changes in subretinal K⁺ concentration occurring upon light stimulation, this thanks to its unusual inverse conductance relation to extracellular K⁺. We demonstrate that inactivating this property by mutation Kir7.1-M125R in mice did not affect retinal response to light stimulus, suggesting that a different channel function must be affected in eye disease caused by mutations of the Kir7.1 gene.

Cancer cachexia is a debilitating syndrome characterized by progressive skeletal muscle wasting and systemic inflammation, primarily observed in patients with advanced-stage cancer. Cachexia severely impacts patients' quality of life and even increases mortality rates; however, effective therapeutic interventions remain elusive. To identify key mediators of muscle atrophy, we integrated more than one hundred bulk and single-cell transcriptomic datasets from diverse murine cachexia models, including colorectal, lung, and pancreatic cancer. This analysis identified leucine-rich alpha-2-glycoprotein 1 (Lrg1), as consistently upregulated in skeletal muscle endothelial cells across cachexia models and progressively increased during disease progression. Functional studies demonstrated that recombinant Lrg1 induced myotube atrophy in vitro, accompanied by reduced fusion index, shortened myotube length, and increased expression of the atrogenes MAFbx and MuRF1. Neutralization of Lrg1 or pharmacological inhibition of Stat3 prevented these effects. Our findings nominate Lrg1 as a candidate biomarker and potential therapeutic target for preventing skeletal muscle wasting in cancer cachexia.

Background: Monocytes and regulatory non-coding RNAs play a crucial role in the development of atherosclerosis (ATH). We have previously shown that miR-125b-5p was upregulated in aortic macrophages and the aim of this paper was to further study the "in vivo" impact of miR-125b-5p in ATH progression. Methods: Eight-weeks-old ApoE^-/- mice, fed with a high-fat diet for 14 weeks, were treated with a miR-125b-5p mimic, with its specific antagonist (antagomiR-125b), with a control scrambled sequence (control oligonucleotide SC) or with a control vehicle with phosphate-buffered saline (PBS) for 4 weeks. Results: Treatment with the miR-125b-5p mimic increased plaque sizes, macrophage infiltration, and NF-κB activation compared to PBS control, independently of cholesterol levels. In contrast, treatment with a specific antagomir produced opposite effects and increased the number of M2 macrophages. Lastly, the miR-125b-5p mimic was found to reduce expression of the chemokine receptor CCR7 in THP-1 cells, in RAW264.7 cells, as well as in the aortas and livers of mice, while the antagomiR-125b increased CCR7 expression. Reduced CCR7 expression was also observed in the aorta of patients with coronary artery disease. Conclusions: MiR-125b-5p mimic increased inflammation and ATH progression. Targeting miR-125b-5p with a specific antagomir reduced plaque size and macrophage infiltration and increased expression of the chemokine receptor CCR7. These results support a role for miR-125b-5p in the upregulation of CCR7 expression and monocyte trafficking, thus restricting vascular inflammation in ATH progression.

After MI, increased spontaneous SR-Ca²⁺-releases depolarize the membrane and trigger action potentials (APs) in cardiac Purkinje cells (Pcells). This abnormal Ca²⁺-activity is involved in ventricular fibrillation. Spontaneous Ca²⁺-transients analysis suggested that intensification of SR-Ca²⁺-uptake accounts for the abnormal SR-Ca²⁺-release in post-MI Pcells. Increased SR-Ca²⁺-pump (SERCA) density, PLB-dependent Ca²⁺-pump activation, and modification of the pump Ca²⁺-transport properties can mediate an increase in SR-Ca²⁺-uptake. We examined whether Pcells of ischemic hearts show signs of these alterations, hence supporting the hypothesis of post-MI increase in SR-Ca²⁺-uptake. Methods. Pcells were prepared from hearts with and without MI in dogs, sheep, pigs, and humans. The distribution of SR-Ca²⁺-pumps and phosphorylated forms of PLB, pPLBSer16 and pPLBThr17, was captured by specific immunofluorescence and confocal microscopy. Protein and transcript levels of SERCA2 sub-isoforms were measured in Purkinje fibers and myocardium by WB and RT-qPCR, respectively. Results. In normal hearts, Ca²⁺-pumps and PLB antibodies co-localized throughout Pcells. After MI, Ca²⁺-pump staining exhibited larger intensity in peripheral compared to central regions of Pcells. Phosphorylated PLB staining was unchanged, indicating no alteration of the pump-β-adrenergic regulation after MI. Expression of the regular cardiac pump, SERCA2a, was preserved. However, the emergence of another pump, SERCA2b, was found after MI. The addition of SERCA2b to the existing SERCA2a expression increased the total pump density, which was consistent with an augmentation of SR-Ca²⁺-uptake in Pcells after MI. Conclusion. After MI, the peripheral region of Pcells seems to express the SERCA2b pump sub-isoform, which is consistent with larger pump density and intensification of SR-Ca²⁺-uptake.

This systematic review investigates the role of non-coding RNAs (ncRNAs), including miRNAs, lncRNAs, circRNAs, and tRNAs, in regulating mitochondrial biogenesis, dynamics, oxidative phosphorylation, and mitophagy in skeletal muscle and the potential applications of these ncRNAs in exercise molecular physiology. We conducted a comprehensive search in PubMed, Scopus, and Web of Science databases, identifying 45 relevant studies out of 2,378 records. The main findings indicate that miRNAs such as miR-128, miR-133a, miR-696, and miR-499 are critical regulators of mitochondrial function. Moreover, lncRNAs (lncEDCH1 and lncRNA-H19) and circRNA (circ-PTPN4) significantly influence mitochondrial biogenesis and function. Exercise interventions were shown to modulate the expression of these ncRNAs, particularly miR-133a and miR-696, leading to enhanced mitochondrial biogenesis and function. The review highlights the potential of these ncRNAs as biomarkers and therapeutic targets for improving mitochondrial function and treating metabolic and mitochondrial disorders. Further research is needed to explore the muscle-specific and exercise-modality-specific effects of ncRNAs to develop personalized interventions. Understanding the complex regulatory mechanisms of ncRNAs in mitochondrial adaptations can pave the way for innovative therapeutic strategies in exercise molecular physiology and metabolic health.

Increased degradation of the endothelial glycocalyx (EGX) is associated with cardiovascular disease. However, whether EGX impairment drives endothelial dysfunction or reflects disease severity remains unclear. Prior studies investigating EGX function primarily used two-dimensional (2D) endothelial cell cultures, which poorly mimic the endothelial microenvironment, particularly lacking luminal shear flow. To address these limitations, we leveraged a three-dimensional (3D) human endothelium-on-a-chip to examine the roles of EGX components, namely heparan sulfate (HS) and sialic acid (SA), in regulating vascular permeability and monocyte adhesion. EGX expression was markedly higher in perfused 3D human umbilical vein endothelial cells (HUVECs) cultures than in 2D cultures. In 3D HUVECs, tumor necrosis factor-alpha, a disruptor of endothelial function, did not reduce EGX expression, whereas dengue nonstructural protein 1 downregulated EGX. In 3D HUVECs, HS degradation by heparinase III significantly increased endothelial permeability to 70-kDa fluorescein isothiocyanate-dextran without inducing cytotoxicity, whereas SA cleavage by neuraminidase reduced vascular permeability. Interestingly, neither HS nor SA cleavage affected 3D human coronary artery endothelial cells (HCAECs) permeability. However, neuraminidase treatment significantly increased monocyte adhesion in both 3D HUVECs and HCAECs, an effect not observed in heparinase III-treated 3D endothelium from either vessel bed. These findings demonstrate that HS and SA play distinct roles in regulating endothelial barrier function and vascular inflammation in 3D human endothelium.