Regulation of specific abnormal calcium signals in the hippocampal CA1 and primary cortex M1 alleviates the progression of temporal lobe epilepsy

Temporal lobe epilepsy is a multifactorial neurological dysfunction syndrome that is refractory,resistant to antiepileptic drugs,and has a high recurrence rate.The pathogenesis of temporal lobe epilepsy is complex and is not fully understood.Intracellular calcium dynamics have been implicated in temporal lobe epilepsy.However,the effect of fluctuating calcium activity in CA1 pyramidal neurons on temporal lobe epilepsy is unknown,and no longitudinal studies have investigated calcium activity in pyramidal neurons in the hippocampal CA1 and primary motor cortex M1 of freely moving mice.In this study,we used a multichannel fiber photometry system to continuously record calcium signals in CA1 and M1 during the temporal lobe epilepsy process.We found that calcium signals varied according to the grade of temporal lobe epilepsy episodes.In particular,cortical spreading depression,which has recently been frequently used to represent the continuously and substantially increased calcium signals,was found to correspond to complex and severe behavioral characteristics of temporal lobe epilepsy ranging from gradeⅡto gradeⅤ.However,vigorous calcium oscillations and highly synchronized calcium signals in CA1 and M1 were strongly related to convulsive motor seizures.Chemogenetic inhibition of pyramidal neurons in CA1 significantly attenuated the amplitudes of the calcium signals corresponding to gradeⅠepisodes.In addition,the latency of cortical spreading depression was prolonged,and the above-mentioned abnormal calcium signals in CA1 and M1 were also significantly reduced.Intriguingly,it was possible to rescue the altered intracellular calcium dynamics.Via simultaneous analysis of calcium signals and epileptic behaviors,we found that the progression of temporal lobe epilepsy was alleviated when specific calcium signals were reduced,and that the end-point behaviors of temporal lobe epilepsy were improved.Our results indicate that the calcium dynamic between CA1 and M1 may reflect specific epileptic behaviors corresponding to different grades.Furthermore,the selective regulation of abnormal calcium signals in CA1 pyramidal neurons appears to effectively alleviate temporal lobe epilepsy,thereby providing a potential molecular mechanism for a new temporal lobe epilepsy diagnosis and treatment strategy.

Suppressing a mitochondrial calcium uniporter activates the calcium signaling pathway and promotes cell elongation in cotton

Mitochondrial calcium uniporter(MCU)is a conserved calcium ion(Ca^(2+))transporter in the mitochondrial inner membrane of eukaryotic cells.How MCU proteins regulate Ca^(2+)flow and modulate plant cell development remain largely unclear.Here,we identified the gene GhMCU4 encoding a MCU protein that negatively regulates plant development and fiber elongation in cotton(Gossypium hirsutum).GhMCU4expressed constitutively in various tissues with the higher transcripts in elongating fiber cells.Knockdown of GhMCU4 in cotton significantly elevated the plant height and root length.The calcium signaling pathway was significantly activated and calcium sensor genes,including Ca^(2+)dependent modulator of interactor of constitutively active ROP(GhCMI1),calmodulin like protein(GhCML46),calciumdependent protein kinases(GhCPKs),calcineurin B-like protein(GhCBLs),and CBL-interacting protein kinases(GhCIPKs),were dramatically upregulated in GhMCU4-silenced plants.Metabolic processes were preferentially enriched,and genes related to regulation of transcription were upregulated in GhMCU4-silenced plants.The contents of Ca^(2+)and H_(2)O_(2)were significantly increased in roots and leaves of GhMCU4-silenced plants.Fiber length and Ca^(2+)and H_(2)O_(2)contents in fibers were significantly increased in GhMCU4-silenced plants.This study indicated that GhMCU4 plays a negative role in regulating cell elongation in cotton,thus expanding understanding in the role of MCU proteins in plant growth and development.

Calmodulins and calmodulin-like proteins-mediated plant organellar calcium signaling networks under abiotic stress

Plant calmodulins(CaMs)and calmodulin-like proteins(CMLs)mediate Ca~(2+)signaling in response to abiotic stresses.Manipulation of this signaling in crops could increase stress tolerance.We review methods for detecting Ca~(2+)signals,regulatory roles of Ca Ms and CMLs,binding targets,and Ca~(2+)networks under abiotic stress in organelles.

Kai-Xin-San regulates synaptic plasticity through calcium signaling to alleviate symptoms of depression-like behavioral disorders in mice

Background:Kai-Xin-San,a classical Chinese medicine prescription,has been widely applied in the clinical therapy for depression,but its pharmacological mechanism remains to be further explored.Based on network pharmacology,molecular docking and animal experiments,the research is performed to exploit pharmacological mechanism of Kai-Xin-San for treating depression.Methods:Obtain chemical components and potential targets of Kai-Xin-San through Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform,Encyclopedia of Traditional Chinese Medicine and Bioinformatics Analysis Tool for Molecular Mechanism of Traditional Chinese Medicine databases,and then screen the active ingredients of each herb in accordance with absorption,distribution,metabolism,and excretion.The GenCards,Online Mendelian Inheritance in Man,Therapeutic Target database and DrugBank databases were used to obtain the major targets of depression,and the STRING platform was used to construct the protein-protein interaction network and explore the potential protein functional modules in the network.The targets were subjected to Gene Ontology enrichment analysis and Kyoto Encyclopedia of Genes and Genomes pathway analysis by STRING database and Metascape database.The interaction network of“Kai-Xin-San active components-depression-targets-pathways”was constructed by Cytoscape,and molecular docking verification was performed by Auto Dock tools.Finally,animal experiments were carried out for further verification.The chronic restraint stress depression model was established and mice were randomly divided into 4 groups:control group,chronic restraint stress group,fluoxetine group and Kai-Xin-San group.Behavioral tests were used to evaluate the depressive phenotype of mice.The expression of CaMKII-,synaptophysin,poststroke depression-95,and CACNA1C were all detected using a western blot.Results:Network analysis shows that Kai-Xin-San may mainly regulate calcium signaling pathway to exert antidepressant effects.A majority of the targets and components have good binding activity,according to the molecular docking studies.In the current study,behavioral tests showed that Kai-Xin-San could effectively alleviate depression-like behaviors in mice compared with the chronic restraint stress group,which effect was comparable to fluoxetine.Meanwhile,compared with the chronic restraint stress group,protein levels of CACNA1C,CaMKII-α,synaptophysin and poststroke depression-95 were significantly increased(P<0.05).Conclusion:The research initially identifies the multi-component,multi-target,and multi-path mechanism of Kai-Xin-San in the treatment of depression.Kai-Xin-San may improve synaptic plasticity through calcium signaling pathway to exert antidepressant effects.

Research process of the calcium signaling in cyanobacteria

The potential involvement of calcium in signalling in cyanobacteria has been investigated in recent years. Enough evidences showed that the cyanobacteria were capable of sensing and distinguishing different environmental stimuli, and making responses in ways of Ca^2＋ transients, which were the results of influx or efflux of Ca^2＋ aroused by different environmental stimuli. The calcium signal elicited by nitrogen starvation was crucial to heterocyst differentiation in filamentous cyanobacteria Anabaena species. Identification of a calcium-binding protein （CcbP） from Anabaena sp. PCC 7120 provided further evidence, and the degradation and down-regulation of CcbP accounted for the generation of calcium signal when nitrogen starvation exits. However, the encoding and decoding mechanisms of the calcium signals in cyanobacteria still remain unclear. In order to reveal the exact role of it, a detailed, systematic investigation will be needed, especially for the calcium dynamics at the single cell level.

Glucocorticoid receptor signaling in the brain and its involvement in cognitive function

The hypothalamic-pituitary-adrenal axis regulates the secretion of glucoco rticoids in response to environmental challenges.In the brain,a nuclear receptor transcription fa ctor,the glucocorticoid recepto r,is an important component of the hypothalamicpituitary-a d renal axis's negative feedback loop and plays a key role in regulating cognitive equilibrium and neuroplasticity.The glucoco rticoid receptor influences cognitive processes,including glutamate neurotransmission,calcium signaling,and the activation of brain-derived neurotrophic factor-mediated pathways,through a combination of genomic and non-genomic mechanisms.Protein interactions within the central nervous system can alter the expression and activity of the glucocorticoid receptor,there by affecting the hypothalamic-pituitary-a d renal axis and stress-related cognitive functions.An appropriate level of glucocorticoid receptor expression can improve cognitive function,while excessive glucocorticoid receptors or long-term exposure to glucoco rticoids may lead to cognitive impairment.Patients with cognitive impairment-associated diseases,such as Alzheimer's disease,aging,depression,Parkinson's disease,Huntington's disease,stroke,and addiction,often present with dysregulation of the hypothalamic-pituitary-adrenal axis and glucocorticoid receptor expression.This review provides a comprehensive overview of the functions of the glucoco rticoid receptor in the hypothalamic-pituitary-a d renal axis and cognitive activities.It emphasizes that appropriate glucocorticoid receptor signaling fa cilitates learning and memory,while its dysregulation can lead to cognitive impairment.This provides clues about how glucocorticoid receptor signaling can be targeted to ove rcome cognitive disability-related disorders.

Pan-cancer analysis of ORAI family proteins and their genomic alterations

Background:The ORAI family of proteins,comprising ORAI1,ORAI2,and ORAI3,plays a crucial role in the regulation of intracellular calcium signaling,which is essential for various cellular functions including proliferation,differentiation,and apoptosis.Dysregulation of calcium signaling has been implicated in cancer pathogenesis,influencing tumor progression,metastasis,and resistance to therapy.This study aims to provide a comprehensive analysis of the ORAI family members across a broad spectrum of cancers.Methods:Publicly available datasets from The Cancer Genome Atlas and the Gene Expression Omnibus were utilized.RNA sequencing data,mutation profiles,copy number variation data,and methylation data across different cancer types were analyzed.Differential expression analysis,survival analysis,copy number variation analysis,mutation analysis,methylation analysis,immune cell infiltration analysis using the Cibersort algorithm,and gene set enrichment analysis were conducted using R software.Results:ORAI1 and ORAI3 were significantly upregulated in glioblastoma multiforme,whereas ORAI2 was notably downregulated in kidney chromophobe and pancreatic adenocarcinoma.ORAI2 exhibited higher mutation rates and copy number gains in multiple cancers compared to ORAI1 and ORAI3.The hypermethylation of ORAI2 in head and neck squamous cell carcinoma,esophageal carcinoma,and glioblastoma multiforme negatively correlated with its gene expression.ORAI1 and ORAI3 expression positively correlated with regulatory T cells infiltration,whereas ORAI2 showed a negative correlation with CD8^(+)T cell infiltration.Gene set enrichment analysis revealed that ORAI1 and ORAI2 are associated with immune-related pathways,while ORAI3 is linked to MYC targets and oxidative phosphorylation.Conclusion:Our pan-cancer analysis reveals significant differential expression,genomic alterations,and epigenetic regulation of ORAI family members across various cancers.ORAI1 and ORAI3 appear to promote an immunosuppressive environment,whereas ORAI2 may function as a tumor suppressor in certain contexts.These findings provide a foundation for future research targeting ORAI-mediated pathways in cancer therapy and highlight the therapeutic potential of ORAI proteins.

Calcium signaling of pancreatic acinar cells in the pathogenesis of pancreatitis

Pancreatitis is an increasingly common and sometimes severe disease that lacks a specific therapy. The pathogenesis of pancreatitis is still not well understood. Calcium (Ca2+) is a versatile carrier of signals regulating many aspects of cellular activity and plays a central role in controlling digestive enzyme secretion in pancreatic acinar cells. Ca2+ overload is a key early event and is crucial in the pathogenesis of many diseases. In pancreatic acinar cells, pathological Ca2+ signaling (stimulated by bile, alcohol metabolites and other causes) is a key contributor to the initiation of cell injury due to prolonged and global Ca2+ elevation that results in trypsin activation, vacuolization and necrosis, all of which are crucial in the development of pancreatitis. Increased release of Ca2+ from stores in the intracellular endoplasmic reticulum and/or increased Ca2+ entry through the plasma membrane are causes of such cell damage. Failed mitochondrial adenosine triphosphate (ATP) production reduces re-uptake and extrusion of Ca2+ by the sarco/endoplasmic reticulum Ca2+-activated ATPase and plasma membrane Ca2+-ATPase pumps, which contribute to Ca2+ overload. Current findings have provided further insight into the roles and mechanisms of abnormal pancreatic acinar Ca2+ signals in pancreatitis. The lack of available specific treatments is therefore an objective of ongoing research. Research is currently underway to establish the mechanisms and interactions of Ca2+ signals in the pathogenesis of pancreatitis.

Calcium Signaling is Involved in Negative Phototropism of Rice Seminal Roots

Calcium ions （Ca2＋） act as an intracellular second messenger and affect nearly all aspects of cellular life. They are functioned by interacting with polar auxin transport, and the negative phototropism of plant roots is caused by the transport of auxin from the irradiated side to the shaded side of the roots. To clarify the role of calcium signaling in the modulation of rice root negative phototropism, as well as the relationship between polar auxin transport and calcium signaling, calcium signaling reagents were used to treat rice seminal roots which were cultivated in hydroculture and unilaterally illuminated at an intensity of 100-200 pmol/（m2.s） for 24 h. Negative phototropism curvature and growth rate of rice roots were both promoted by exogenous CaCI2 lower than 100 pmol/L, but inhibited by calcium channel blockers （verapamil and LaCI3）, calcineurin inhibitor （chlorpromazine, CPZ）, and polar auxin transport inhibitor （N-l-naphthylphthalamic acid, NPA）. Roots stopped growing and negative phototropism disappeared when the concentrations increased to 100 pmol/L verapamil, 12.500 ~Jmol/L LaCI3, 60 pmol/L CPZ, and 6 pmol/L NPA. Moreover, 100 pmol/L CaCI2 could relieve the inhibition of LaCI3, verapamil and NPA. The enhanced negative phototropism curvature was caused by the transportation of more auxin from the irradiated side to the shaded side in the presence of exogenous Ca2＋. Calcium signaling plays a key role as a second messenger in the process of light signal regulation of rice root growth and negative phototropism.

Connecting Calcium-Based Nanomaterials and Cancer:From Diagnosis to Therapy

As the indispensable second cellular messenger,calcium signaling is involved in the regulation of almost all physiological processes by activating specific target proteins.The importance of calcium ions(Ca^(2+))makes its“Janus nature”strictly regulated by its concentration.Abnormal regulation of calcium signals may cause some diseases;however,artificial regulation of calcium homeostasis in local lesions may also play a therapeutic role.“Calcium overload,”for example,is characterized by excessive enrichment of intracellular Ca^(2+),which irreversibly switches calcium signaling from“positive regulation”to“reverse destruction,”leading to cell death.However,this undesirable death could be defined as“calcicoptosis”to offer a novel approach for cancer treatment.Indeed,Ca^(2+)is involved in various cancer diagnostic and therapeutic events,including calcium overload-induced calcium homeostasis disorder,calcium channels dysregulation,mitochondrial dysfunction,calcium-associated immunoregulation,cell/vascular/tumor calcification,and calcification-mediated CT imaging.In paral-lel,the development of multifunctional calcium-based nanomaterials(e.g.,calcium phosphate,calcium carbonate,calcium peroxide,and hydroxyapatite)is becoming abundantly available.This review will highlight the latest insights of the calcium-based nanomaterials,explain their application,and provide novel perspective.Identifying and characterizing new patterns of calcium-dependent signaling and exploiting the disease element linkage offer additional translational opportunities for cancer theranostics.

Role of calcium in polycystic kidney disease:From signaling to pathology

Autosomal dominant polycystic kidney disease （ADPKD） is the most common inherited monogenic kidney disease. Characterized by the development and growth of cysts that cause progressive kidney enlargement, it ultimately leads to end-stage renal disease. Approximately 85% of ADPKD cases are caused by mutations in the PKD1 gene, while mutations in the PKD2 gene account for the remaining 15% of cases. The PKD1 gene encodes for polycystin-1 （PC1）, a large multi-functional memb-rane receptor protein able to regulate ion channel complexes, whereas polycystin-2 （PC2）, encoded by the PKD2 gene, is an integral membrane protein that functions as a calcium-permeable cation channel, located mainly in the endoplasmic reticulum （ER）. In the primary cilia of the epithelial cells, PC1 interacts with PC2 to form a polycystin complex that acts as a mechanosensor, regulating signaling pathways involved in the differentiation of kidney tubular epithelial cells. Despite progress in understanding the function of these proteins, the molecular mechanisms associated with the pathogenesis of ADPKD remain unclear. In this review we discuss how an imbalance between functional PC1 and PC2 proteins may disrupt calcium channel activities in the cilium, plasma membrane and ER, thereby altering intracellular calcium signaling and leading to the aberrant cell proliferation and apoptosis associated with the development and growth of renal cysts. Research in this feld could lead to the discovery of new molecules able to rebalance intracellular calcium, thereby normalizing cell proliferation and reducing kidney cyst progression.

T-type calcium channel expression in cultured human neuroblastoma cells

Human neuroblastoma cells （SH-SY5Y） have similar structures and functions as neural cells and have been frequently used for cell culture studies of neural cell functions.Previous studies have revealed L-and N-type calcium channels in SH-SY5Y cells.However,the distribution of the low-voltage activated calcium channel （namely called T-type calcium channel,including Cav3.1,Cav3.2,and Cav3.3） in SH-SY5Y cells remains poorly understood.The present study detected mRNA and protein expres-sion of the T-type calcium channel （Cav3.1,Cav3.2,and Cav3.3） in cultured SH-SY5Y cells using real-time polymerase chain reaction （PCR） and western blot analysis.Results revealed mRNA and protein expression from all three T-type calcium channel subtypes in SH-SY5Y cells.Moreover,Cav3.1 was the predominant T-type calcium channel subtype in SH-SY5Y cells.

Intracellular calcium ions facilitate dengue virus entry into endothelial cells and compromise endothelial barrier integrity

Objective:To investigate the involvement of Ca^(2+)in dengue virus(DENV)-infected human umbilical vein endothelial cells(HUVECs)and the disruption of endothelial integrity.Methods:HUVECs were infected with DENV-2 in the presence of intracellular Ca^(2+)or endoplasmic reticulum Ca^(2+)chelators.Virus infectivity was measured by focus-forming assay and quantitative RT-PCR.Intracellular Ca^(2+)was measured using Fluo-4-AM dye.VE-cadherin and focal adhesion kinase(FAK)expressions were investigated by immunofluorescence and immunoblotting assays,respectively.Results:DENV infection increased intracellular cytosolic Ca^(2+)levels and caused disassembly of the adherens junction protein,VEcadherin as evidenced by decreased VE-cadherin expression at the periphery of DENV-2 infected HUVECs.Depletion of intracellular Ca^(2+)stores,particularly those of the endoplasmic reticulum Ca^(2+),significantly decreased DENV yield in HUVECs.Decreased virus yield following the depletion of intracellular Ca^(2+)was caused by the inhibition of viral entry into HUVECs and not the inhibition of viral binding or attachment.DENV-2 infection also resulted in Ca^(2+)-dependent activation of FAK.Conclusions:Intracellular Ca^(2+)is required for the early phases of DENV infection in endothelial cells.Increased cytosolic Ca^(2+)levels in endothelial cells during DENV infection activated FAK,disrupted adherens junctions and compromised barrier integrity.Thus,Ca^(2+)plays an important role in DENV infection in endothelial cells.

ICAM-1 depletion in the center of immunological synapses is important for calcium releasing in T-cells

T-cell activation requires the formation of the immunological sy napse(IS)bet ween a T-cll and anantigen-presenting cell(AP C)to control the development of the adaptive immune response.How-ever,calcium release,an initial signal of T-cell activation,has been found to occur before IS for-mation.The mechanism for triggering the calcium signaling and relationship bet ween calciumrelease and IS format ion remains unclear.Herein,using live-cell imaging,we found that int ercellularadhesion molecule 1(ICAM-1),an essential mdlecule for IS formation,accumulated and then wasdepleted at the center of the synapse before complete IS formation.During the proces of ICAM1depletion,calcium was released.if ICAM-1 failed to be depleted from the center of the synapse,thesustained calcium signaling could not be induced.Moreover,depletion of ICAM-1 in ISs preferen-tially ccurred with the contact of antigen-specific T-cels and dendritic clls(DCs).Blocking thebinding ofICA M-1 and lymphocy te finction-associated antigen 1(LFA-1),ICAM-1 failed to depleteat the center of the synapse,and calcium release in T-clls decreased.In studying the mechanism ofhow the depletion ofiCA M1 could influence calcium release in T-clls,we found that the movementof ICAM-1 was associat ed with the localization of LFA-1 in the IS,which afected the localization ofcalcium microdomains,ORAIl and mitochondria in IS.Therefore,the depletion of ICAM-1 in the center of the synapse is an important factor for an initial sust ained calcium release in T-cells.

Effects of Propofol on Glutamate-Induced Calcium Mobilization in Presynaptic Boutons of Rat Hippocampal Neurons

Recent reports have suggested that various general anesthetics affect presynaptic processes in the central nervous system. However, characterizations of the influence of intravenous anesthetics on neurotransmitter release from presynaptic nerve terminals (boutons) are insufficient. Because the presynaptic calcium concentration ([Ca2+]pre) regulates neurotransmitter release, we investigate the effects of the intravenous anesthetic propofol on neurotransmitter release by measuring [Ca2+]pre in the presynaptic boutons of individual dissociated hippocampal neurons. Brain slices were prepared from Sprague–Dawley rats (10 - 14 days of age). The hippocampal CA1 area was isolated with a fire-polished glass pipette, which vibrated horizontally to dissociate hippocampal CA1 neurons along with their attached presynaptic boutons. Presynaptic boutons were visualized under a confocal laser scanning microscope after staining with FM1-43 dye, and [Ca2+]pre was measured using fluo-3 AM dye. Glutamate (3 – 100 μM) administration increased [Ca2+]pre in Ca2+- containing external solution in a concentration-dependent manner. Propofol (3 – 30 μM) dose-dependently suppressed this glutamate (30 μM)-induced increase in [Ca2+]pre in boutons attached to dendrites, but not to the soma or base of the dendritic tree. The large majority of excitatory synapses on CA1 neurons are located on dendritic spines;therefore, propofol may affect glutamate-induced Ca2+ mobilization in excitatory, but not inhibitory, presynaptic boutons. Propofol may possibly have some effect on glutamate-regulated neurotransmitter release from excitatory presynaptic nerve terminals through inhibiting the increase in [Ca2+]pre induced by glutamate.

From static to dynamic:live observation of the support system after ischemic stroke by two photon-excited fluorescence laser-scanning microscopy

Ischemic stroke is one of the most common causes of mortality and disability worldwide.However,treatment efficacy and the progress of research remain unsatisfactory.As the critical support system and essential components in neurovascular units,glial cells and blood vessels(including the bloodbrain barrier)together maintain an optimal microenvironment for neuronal function.They provide nutrients,regulate neuronal excitability,and prevent harmful substances from entering brain tissue.The highly dynamic networks of this support system play an essential role in ischemic stroke through processes including brain homeostasis,supporting neuronal function,and reacting to injuries.However,most studies have focused on postmortem animals,which inevitably lack critical information about the dynamic changes that occur after ischemic stroke.Therefore,a high-precision technique for research in living animals is urgently needed.Two-photon fluorescence laser-scanning microscopy is a powerful imaging technique that can facilitate live imaging at high spatiotemporal resolutions.Twophoton fluorescence laser-scanning microscopy can provide images of the whole-cortex vascular 3D structure,information on multicellular component interactions,and provide images of structure and function in the cranial window.This technique shifts the existing research paradigm from static to dynamic,from flat to stereoscopic,and from single-cell function to multicellular intercommunication,thus providing direct and reliable evidence to identify the pathophysiological mechanisms following ischemic stroke in an intact brain.In this review,we discuss exciting findings from research on the support system after ischemic stroke using two-photon fluorescence laser-scanning microscopy,highlighting the importance of dynamic observations of cellular behavior and interactions in the networks of the brain’s support systems.We show the excellent application prospects and advantages of two-photon fluorescence laser-scanning microscopy and predict future research developments and directions in the study of ischemic stroke.

Immortalized hippocampal astrocytes from 3xTg-AD mice,a new model to study disease-related astrocytic dysfunction:a comparative review

Alzheimer's disease(AD)is characterized by complex etiology,long-lasting pathogenesis,and celltype-specific alterations.Currently,there is no cure for AD,emphasizing the urgent need for a comprehensive understanding of cell-specific pathology.Astrocytes,principal homeostatic cells of the central nervous system,are key players in the pathogenesis of neurodegenerative diseases,including AD.Cellular models greatly facilitate the investigation of cell-specific pathological alterations and the dissection of molecular mechanisms and pathways.Tumor-derived and immortalized astrocytic cell lines,alongside the emerging technology of adult induced pluripotent stem cells,are widely used to study cellular dysfunction in AD.Surprisingly,no stable cell lines were available from genetic mouse AD models.Recently,we established immortalized hippocampal astroglial cell lines from amyloid-βprecursor protein/presenilin-1/Tau triple-transgenic(3xTg)-AD mice(denominated as wild type(WT)-and 3Tg-iAstro cells)using retrovirus-mediated transduction of simian virus 40 large T-antigen and propagation without clonal selection,thereby maintaining natural heterogeneity of primary cultures.Several groups have successfully used 3Tg-iAstro cells for single-cell and omics approaches to study astrocytic AD-related alterations of calcium signaling,mitochondrial dysfunctions,disproteostasis,altered homeostatic and signaling support to neurons,and blood-brain barrier models.Here we provide a comparative overview of the most used models to study astrocytes in vitro,such as primary culture,tumor-derived cell lines,immortalized astroglial cell lines,and induced pluripotent stem cell-derived astrocytes.We conclude that immortalized WT-and 3Tg-iAstro cells provide a noncompetitive but complementary,low-cost,easy-to-handle,and versatile cellular model for dissection of astrocyte-specific AD-related alterations and preclinical drug discovery.

The link between intracellular calcium signaling and exosomal PD-L1 in cancer progression and immunotherapy

Exosomes are small membrane vesicles containing microRNA,RNA,DNA fragments,and proteins that are transferred from donor cells to recipient cells.Tumor cells release exo-somes to reprogram the factors associated with the tumor microenvironment(TME)causing tu-mor metastasis and immune escape.Emerging evidence revealed that cancer cell-derived exosomes carry immune inhibitory molecule program death ligand 1(PD-L1)that binds with re-ceptor program death protein 1(PD-1)and promote tumor progression by escaping immune response.Currently,some FDA-approved monoclonal antibodies are clinicallyused for cancer treatment by blocking PD-1/PD-L1 interaction.Despite notable treatment outcomes,some pa-tients show poor drug response.Exosomal PD-L1 plays a vital role in lowering the treatment response,showing resistance to PD-1/PD-L1 blockage therapy through recapitulating the ef-fect of cell surface PD-L1.To enhance therapeutic response,inhibition of exosomal PD-L1 is required.Calcium signaling is the central regulator of tumorigenesis and can regulate exosome biogenesis and secretion by modulating Rab GTPase family and membrane fusion factors.Im-mune checkpoints are also connected with calcium signaling and calcium channel blockers like amlodipine,nifedipine,lercanidipine,diltiazem,and verapamil were also reported to suppress cellular PD-L1 expression.Therefore,to enhance the PD-1/PD-L1 blockage therapy response,the reduction of exosomal PD-L1 secretion from cancer cells is in our therapeutic consider-ation.In this review,we proposed a therapeutic strategy by targeting calcium signaling to inhibit the expression of PD-L1-containing exosome levels that could reduce the anti-PD-1/PD-L1 therapy resistance and increase the patient's drug response rate.

Molecular regulation of calcium-sensing receptor(CaSR)-mediated signaling

Calcium-sensing receptor(CaSR),a family C G-protein-coupled receptor,plays a crucial role in regulating calcium homeostasis by sensing small concentration changes of extracellular Ca^(2+),Mg^(2+),amino acids(e.g.,L-Trp and L-Phe),small peptides,anions(e.g.,HCO_(3)^(-)and PO_(4)^(3-)),and pH.CaSR-mediated intracellular Ca^(2+)signaling regulates a diverse set of cellular processes including gene transcription,cell proliferation,differentiation,apoptosis,muscle contraction,and neuronal transmission.Dysfunction of CaSR with mutations results in diseases such as autosomal dominant hypocalcemia,familial hypocalciuric hypercalcemia,and neonatal severe hyperparathyroidism.CaSR also influences calciotropic disorders,such as osteoporosis,and noncalciotropic disorders,such as cancer,Alzheimer's disease,and pulmonary arterial hypertension.This study first reviews recent advances in biochemical and structural determination of the framework of CaSR and its interaction sites with natural ligands,as well as exogenous positive allosteric modulators and negative allosteric modulators.The establishment of the first CaSR protein-protein interactome network revealed 94 novel players involved in protein processing in endoplasmic reticulum,trafficking,cell surface expression,endocytosis,degradation,and signaling pathways.The roles of these proteins in Ca^(2+)-dependent cellular physiological processes and in CaSR-dependent cellular signaling provide new insights into the molecular basis of diseases caused by CaSR mutations and dysregulated CaSR activity caused by its protein interactors and facilitate the design of therapeutic agents that target CaSR and other family C G-protein-coupled receptors.

Molecular Mechanism Underlying Plant Response to Cold Stress

Low temperature stress is one of the most important factors limiting plant growth and geographical distribution.In order to adapt to low temperature,plants have evolved strategies to acquire cold tolerance,known as,cold acclimation.Current molecular and genomic studies have reported that annual herbaceous and perennial woody plants share similar cold acclimation mechanisms.However,woody perennials also require extra resilience to survive cold winters.Thus,trees have acquired complex dynamic processes to control the development of dormancy and cold resistance,ensuring successful tolerance during the coldest winter season.In this review,we systemically described how woody plants perceive and transduce cold stress signals through a series of physiological changes such as calcium signaling,membrane lipid,and antioxidant changes altering downstream gene expression and epigenetic modification,ultimately bud dormancy.We extended the discussion and reviewed the processes endogenous phytohormones play in regulating the cold stress.We believe that this review will aid in the comprehension of underlying mechanisms in plant acclimation to cold stress.