Search

Polyamines are molecules with multiple amino groups that are essential for cellular function. The major polyamines are putrescine, spermidine, spermine, and cadaverine. Polyamines are important for posttranscriptional regulation, autophagy, programmed cell death, proliferation, redox homeostasis, and ion channel function. Their levels are tightly controlled. High levels of polyamines are associated with proliferative pathologies such as cancer, whereas low polyamine levels are observed in aging, and elevated polyamine turnover enhances oxidative stress. Polyamine metabolism is implicated in several pathophysiological processes in the nervous, immune, and cardiovascular systems. Currently, manipulating polyamine levels is under investigation as a potential preventive treatment for several pathologies, including aging, ischemia/reperfusion injury, pulmonary hypertension, and cancer. Although polyamines have been implicated in many intracellular mechanisms, our understanding of these processes remains incomplete and is a topic of ongoing investigation. Here, we discuss the regulation and cellular functions of polyamines, their role in physiology and pathology, and emphasize the current gaps in knowledge and potential future research directions. [ABSTRACT FROM AUTHOR]

The article titled "From promise to success: AJP-Cell Physiology's journey of accomplishments and future vision" discusses the achievements and future plans of the American Journal of Physiology: Cell Physiology. The journal has successfully expanded its content by introducing new topics and collaborating with guest editors. It has also increased its visibility across medical disciplines and attracted submissions from clinicians focused on basic science. The journal plans to further improve by focusing on cutting-edge scientific advancements, implementing a fast-track review process, and increasing diversity of submissions. The article also discusses strategies such as attending conferences, increasing communication with editors and members, and transitioning to an open-access publication model. The author expresses gratitude to the committee, editors, and staff for their contributions. [Extracted from the article]

The American Journal of Physiology: Cell Physiology has experienced an increase in its citation metrics, such as Impact Factor, Cite Score, and h5-index. To attract more authors and readers, the journal has implemented new initiatives, including addressing the challenge of combining rapid publication with rigorous peer review. They have introduced a Fast Track option for high-quality research articles and have launched Calls for Papers to increase diversity and stay updated with advancements in the field. The text references various articles published in the journal, covering topics like matrix homeostasis, renal and cardiac protection, fibrosis, extracellular vesicles, ketone ester administration, ion channels in the musculoskeletal system, and bioreactors for cell culture. The authors express their appreciation to the editorial board and reviewers for their contributions and assure authors of their commitment to maintaining a high-quality publication process. [Extracted from the article]

The extracellular matrix (ECM) is central to the physiology of animal tissues, through its multifaceted roles in tissue structure, mechanical properties, and cell interactions, and by its cell-signaling activities that regulate cell phenotype and behavior. The secretion of ECM proteins typically involves multiple transport and processing steps within the endoplasmic reticulum and the subsequent compartments of the secretory pathway. Many ECM proteins are substituted with various posttranslational modifications (PTMs) and there is increasing evidence of how PTM additions are required for ECM protein secretion or functionality within the extracellular milieu. The targeting of PTM-addition steps may thus offer opportunities to manipulate ECM quality or quantity, in vitro or in vivo. This review discusses selected examples of PTMs of ECM proteins for which the PTM has known importance for anterograde trafficking and secretion of the core protein, and/or loss-of-function of the respectively modifying enzyme leads to alterations of ECM structure or function with pathophysiological consequences in humans. Members of the protein disulfide isomerase (PDI) family have central roles in disulfide bond formation and isomerization within the endoplasmic reticulum, and are discussed in relation to emerging knowledge of the roles of certain PDIs in ECM production in the pathophysiological context of breast cancer. Cumulative data suggest the possible applicability of inhibition of PDIA3 activity to modulate ECM composition and functionality within the tumor microenvironment. [ABSTRACT FROM AUTHOR]

Circadian rhythms in physiology and behavior allow organisms to anticipate the daily environmental changes imposed by the rotation of our planet around its axis. Although these rhythms eventually manifest at the organismal level, a cellular basis for circadian rhythms has been demonstrated. Significant contributors to these cell-autonomous rhythms are daily cycles in gene expression and protein translation. However, recent data revealed cellular rhythms in other biological processes, including ionic currents, ion transport, and cytosolic ion abundance. Circadian rhythms in ion currents sustain circadian variation in action potential firing rate, which coordinates neuronal behavior and activity. Circadian regulation of metal ions abundance and dynamics is implicated in distinct cellular processes, from protein translation to membrane activity and osmotic homeostasis. In turn, studies showed that manipulating ion abundance affects the expression of core clock genes and proteins, suggestive of a close interplay. However, the relationship between gene expression cycles, ion dynamics, and cellular function is still poorly characterized. In this review, I will discuss the mechanisms that generate ion rhythms, the cellular functions they govern, and how they feed back to regulate the core clock machinery. [ABSTRACT FROM AUTHOR]

Adenosine deaminases acting on RNAs convert adenosines (A) to inosines (I) in structured or double-stranded RNAs. In mammals, this process is widespread. In the human transcriptome, more than a million different sites have been identified that undergo an ADAR-mediated A-to-I exchange Inosines have an altered base pairing potential due to the missing amino group when compared to the original adenosine. Consequently, inosines prefer to base pair with cytosines but can also base pair with uracil or adenine. This altered base pairing potential not only affects protein decoding at the ribosome but also influences the folding of RNAs and the proteins that can associate with it. Consequently, an A to I exchange can also affect RNA processing and turnover (Nishikura K. Annu Rev Biochem 79: 321-349, 2010; Brümmer A, Yang Y, Chan TW, Xiao X. Nat Commun 8: 1255, 2017). All of these events will interfere with gene expression and therefore, can also affect cellular and organismic physiology. As double-stranded RNAs are a hallmark of viral pathogens RNA-editing not only affects RNA-processing, coding, and gene expression but also controls the antiviral response to double-stranded RNAs. Most interestingly, recent advances in our understanding of ADAR enzymes reveal multiple layers of regulation by which ADARs can control antiviral programs. In this review, we focus on the recoding of mRNAs where the altered translation products lead to physiological changes. We also address recent advances in our understanding of the multiple layers of antiviral responses and innate immune modulations mediated by ADAR1. [ABSTRACT FROM AUTHOR]

The liver holds central roles in detoxification, energy metabolism, and whole body homeostasis but can develop malignant phenotypes when being chronically overwhelmed with fatty acids and glucose. The global rise of metabolic dysfunctionassociated fatty liver disease (MAFLD) is already affecting a quarter of the global population. Pharmaceutical treatment options against different stages of MAFLD do not yet exist, and several clinical trials against hepatic transcription factors and other proteins have failed. However, emerging roles of noncoding RNAs, including long (lncRNA) and short noncoding RNAs (sRNA), in various cellular processes pose exciting new avenues for treatment interventions. Actions of noncoding RNAs mostly rely on interactions with proteins, whereby the noncoding RNA fine-tunes protein function in a process termed riboregulation. The developmental stage-, disease stage-, and cell type-specific nature of noncoding RNAs harbors enormous potential to precisely target certain cellular pathways in a spatiotemporally defined manner. Proteins interacting with RNAs can be categorized into canonical or noncanonical RNA-binding proteins (RBPs) depending on the existence of classical RNA-binding domains. Both, RNA-and RBP-centric methods have generated new knowledge of the RNA-RBP interface and added an additional regulatory layer. In this review, we summarize recent advances in how RBP-lncRNA interactions and various sRNAs shape cellular physiology and the development of liver diseases such as MAFLD and hepatocellular carcinoma. [ABSTRACT FROM AUTHOR]

Renal blood flow represents >20% of total cardiac output and with this comes the great responsibility of maintaining homeostasis through the intricate regulation of solute handling. Through the processes of filtration, reabsorption, and secretion, the kidneys ensure that solutes and other small molecules are either returned to circulation, catabolized within renal epithelial cells, or excreted through the process of urination. Although this occurs throughout the renal nephron, one segment is tasked with the bulk of solute reabsorption-the proximal tubule. Among others, the renal proximal tubule is entirely responsible for the reabsorption of glucose, a critical source of energy that fuels the body. In addition, it is the only other site of gluconeogenesis outside of the liver. When these processes go awry, pathophysiological conditions such as diabetes and acidosis result. In this review, we highlight the recent advances made in understanding these processes that occur within the renal proximal tubule. We focus on the physiological mechanisms at play regarding glucose reabsorption and glucose metabolism, emphasize the conditions that occur under diseased states, and explore the emerging class of therapeutics that are responsible for restoring homeostasis. [ABSTRACT FROM AUTHOR]

SPOCK1, 2, and 3 are considered matricellular proteoglycans without a structural role. Their functions are only partly elucidated. SPOCK1 was detected in the brain as a member of the neural synapses, then in the neuromuscular junctions. It plays a role in the regulation of the blood-brain barrier. Its best-characterized activity was its oncogenic potential discovered in 2012. Its deleterious effect on tumor progression was detected on 36 different types of tumors by the end of 2020. However, its mode of action is still not completely understood. Furthermore, even less was discovered about its physiological function. The fact that it was found to localize in the mitochondria and interfered with the lipid metabolism indicated that the full discovery of SPOCK1 is still waiting for us. [ABSTRACT FROM AUTHOR]

The importance of defining sex differences across various biological and physiological mechanisms is more pervasive now than it has been over the past 15-20 years. As the muscle biology field pushes to identify small molecules and interventions to prevent, attenuate, or even reverse muscle wasting, we must consider the effect of sex as a biological variable. It should not be assumed that a therapeutic will affect males and females with equal efficacy or equivalent target affinities under conditions where muscle wasting is observed. With that said, it is not surprising to find that we have an unclear or even a poor understanding of the effects of sex or sex hormones on muscle wasting conditions. Although recent investigations are beginning to establish experimental approaches that will allow investigators to assess the impact of sex-specific hormones on muscle wasting, the field still needs rigorous scientific tools that will allow the community to address critical hypotheses centered around sex hormones. The focus of this review is on female sex hormones, specifically estrogens, and the roles that these hormones and their receptors play in skeletal muscle wasting conditions. With the overall review goal of assembling the current knowledge in the area of sexual dimorphism driven by estrogens with an effort to provide insights to interested physiologists on necessary considerations when trying to assess models for potential sex differences in cellular and molecular mechanisms of muscle wasting. [ABSTRACT FROM AUTHOR]

The resolution of inflammation is closely linked with tissue repair. Recent studies have revealed that macrophages suppress inflammatory reactions by producing lipid mediators, called specialized proresolving mediators (SPMs); however, the biological significance of SPMs in tissue repair remains to be fully elucidated in the heart. In this study, we focused on maresin-1 (MaR1) and examined the reparative effects of MaR1 in cardiomyocytes. The treatment with MaR1 increased cell size in cultured neonatal rat cardiomyocytes. Since the expression of fetal cardiac genes was unchanged by MaR1, physiological hypertrophy was induced by MaR1. SR3335, an inhibitor of retinoic acid-related orphan receptor α (RORα), mitigated MaR1-induced cardiomyocyte hypertrophy, consistent with the recent report that RORα is one of MaR1 receptors. Importantly, in response to MaR1, cardiomyocytes produced IGF-1 via RORα. Moreover, MaR1 activated phosphoinositide 3-kinase (PI3K)/Akt signaling pathway and wortmannin, a PI3K inhibitor, or triciribine, an Akt inhibitor, abrogated MaR1-induced cardiomyocyte hypertrophy. Finally, the blockade of IGF-1 receptor by NVP-AEW541 inhibited MaR-1-induced cardiomyocyte hypertrophy as well as the activation of PI3K/Akt pathway. These data indicate that MaR1 induces cardiomyocyte hypertrophy through RORα/IGF-1/PI3K/Akt pathway. Considering that MaR1 is a potent resolving factor, MaR1 could be a key mediator that orchestrates the resolution of inflammation with myocardial repair. [ABSTRACT FROM AUTHOR]

Oogenesis is a complex process resulting in the production of a truly remarkable cell--the oocyte. Oocytes execute many unique processes and functions such as meiotic segregation of maternal genetic material, and essential life-generating functions after fertilization including posttranscriptional support of essential homeostatic and metabolic processes, and activation and reprogramming of the embryonic genome. An essential goal for understanding female fertility and infertility in mammals is to discover critical features driving the production of quality oocytes, particularly the complex regulation of oocyte maternal mRNAs. We report here the first in-depth meta-analysis of oocyte maturation-associated transcriptome changes, using eight datasets encompassing 94 RNAseq libraries for human, rhesus monkey, mouse, and cow. A majority of maternal mRNAs are regulated in a species-restricted manner, highlighting considerable divergence in oocyte transcriptome handling during maturation. We identified 121 mRNAs changing in relative abundance similarly across all four species (92 of high homology), and 993 (670 high homology) mRNAs regulated similarly in at least three of the four species, corresponding to just 0.84% and 6.9% of mRNAs analyzed. Ingenuity Pathway Analysis (IPA) revealed an association of these shared mRNAs with many shared pathways and functions, most prominently oxidative phosphorylation and mitochondrial function. These shared functions were reinforced further by primate-specific and species-specific differentially expressed genes (DEGs). Thus, correct downregulation of mRNAs related to oxidative phosphorylation and mitochondrial function is a major shared feature of mammalian oocyte maturation. [ABSTRACT FROM AUTHOR]

For about half a century, the pharmacology of electroneutral cation-chloride cotransporters has been dominated by a few drugs that are widely used in clinical medicine. Because these diuretic drugs are so good at what they do, there has been little incentive in expanding their pharmacology. The increasing realization that cation-chloride cotransporters are involved in many other key physiological processes and the knowledge that different tissues express homologous proteins with matching transport functions have rekindled interest in drug discovery. This review summarizes the methods available to assess the function of these transporters and describe the multiple efforts that have made to identify new compounds. We describe multiple screens targeting KCC2 function and one screen designed to find compounds that discriminate between NKCC1 and NKCC2. Two of the KCC2 screens identified new inhibitors that are 3-4 orders of magnitude more potent than furosemide. Additional screens identified compounds that purportedly increase cell surface expression of the cotransporter, as well as several FDA-approved drugs that increase KCC2 transcription and expression. The technical details of each screen biased them toward specific processes in the life cycle of the transporter, making these efforts independent and complementary. In addition, each drug discovery effort contributes to our understanding of the biology of the cotransporters. [ABSTRACT FROM AUTHOR]

One of the major roles of the intracardiac nervous system (ICNS) is to act as the final site of signal integration for efferent information destined for the myocardium to enable local control of heart rate and rhythm. Multiple subtypes of neurons exist in the ICNS where they are organized into clusters termed ganglionated plexi (GP). The majority of cells in the ICNS are actually glial cells; however, despite this, ICNS glial cells have received little attention to date. In the central nervous system, where glial cell function has been widely studied, glia are no longer viewed simply as supportive cells but rather have been shown to play an active role in modulating neuronal excitability and synaptic plasticity. Pioneering studies have demonstrated that in addition to glia within the brain stem, glial cells within multiple autonomic ganglia in the peripheral nervous system, including the ICNS, can also act to modulate cardiovascular function. Clinically, patients with atrial fibrillation (AF) undergoing catheter ablation show high plasma levels of S100B, a protein produced by cardiac glial cells, correlated with decreased AF recurrence. Interestingly, S100B also alters GP neuron excitability and neurite outgrowth in the ICNS. These studies highlight the importance of understanding how glial cells can affect the heart by modulating GP neuron activity or synaptic inputs. Here, we review studies investigating glia both in the central and peripheral nervous systems to discuss the potential role of glia in controlling cardiac function in health and disease, paying particular attention to the glial cells of the ICNS. [ABSTRACT FROM AUTHOR]

Is REDD1 a metabolic double agent? Lessons from physiology and pathology. Am J Physiol Cell Physiol 319: C807-C824, 2020. First published September 2, 2020; doi:10.1152/ ajpcell.00340.2020.--The Akt/mechanistic target of rapamycin (mTOR) signaling pathway governs macromolecule synthesis, cell growth, and metabolism in response to nutrients and growth factors. Regulated in development and DNA damage response (REDD)1 is a conserved and ubiquitous protein, which is transiently induced in response to multiple stimuli. Acting like an endogenous inhibitor of the Akt/mTOR signaling pathway, REDD1 protein has been shown to regulate cell growth, mitochondrial function, oxidative stress, and apoptosis. Recent studies also indicate that timely REDD1 expression limits Akt/mTORdependent synthesis processes to spare energy during metabolic stresses, avoiding energy collapse and detrimental consequences. In contrast to this beneficial role for metabolic adaptation, REDD1 chronic expression appears involved in the pathogenesis of several diseases. Indeed, REDD1 expression is found as an early biomarker in many pathologies including inflammatory diseases, cancer, neurodegenerative disorders, depression, diabetes, and obesity. Moreover, prolonged REDD1 expression is associated with cell apoptosis, excessive reactive oxygen species (ROS) production, and inflammation activation leading to tissue damage. In this review, we decipher several mechanisms that make REDD1 a likely metabolic double agent depending on its duration of expression in different physiological and pathological contexts. We also discuss the role played by REDD1 in the cross talk between the Akt/mTOR signaling pathway and the energetic metabolism. [ABSTRACT FROM AUTHOR]

More than a cell biosensor: aryl hydrocarbon receptor at the intersection of physiology and inflammation. Am J Physiol Cell Physiol 318: C1078-C1082, 2020. First published March 25, 2020; doi:10.1152/ajpcell.00493.2019.-- Aryl hydrocarbon receptor (AhR), a highly conserved intracellular transcription factor, is activated by a plethora of ligands of both exogenous and endogenous nature. Besides activating xenobioticmetabolizing enzymes, it is involved in the differentiation and development of hematopoietic, hepatic, nervous and immune systems. More and more data describe its role in the regulation of immune responses and in the onset and progression of inflammation. Particularly, established results view AhR as a downstream target of inflammatory molecules, since its transcription is regulated by the inflammatory cascade. Interleukin 6 (IL-6) has been described to sustain early stages of inflammation and to influence the expression of AhR either directly, following signal transducer and activator of transcription 3 (STAT3) activation, or in combination with other inflammatory mediators, e.g., transforming growth factor-β (TGF-β). In selected inflammatory milieus, once activated, AhR interacts with its targets including the IL-6 promoter, thus originating an autoinflammatory loop. This perspective review brings together evidence that, in some IL-6-driven pathways, AhR is a downstream target that amplifies the duration and extent of inflammation. Considering that many inflammatory mediators can also trigger the activities of AhR as biosensor and activator of xenobiotics metabolism, this issue is of pivotal importance. The individual susceptibly to some environmental ligands of AhR can be probably explained by considering the individual inflammatory state, which could additionally fuel the proinflammatory activity of AhR. Thus, AhR could be considered a transductor of a dynamic, bidirectional connection between internal and external environmental stimuli and the inflammatory response. [ABSTRACT FROM AUTHOR]

Many different subpopulations of subcellular extracellular vesicles (EVs) have been described. EVs are released from all cell types and have been shown to regulate normal physiological homeostasis, as well as pathological states by influencing cell proliferation, differentiation, organ homing, injury and recovery, as well as disease progression. In this review, we focus on the bidirectional actions of vesicles from normal and diseased cells on normal or leukemic target cells; and on the leukemic microenvironment as a whole. EVs from human bone marrow mesenchymal stem cells (MSC) can have a healing effect, reversing the malignant phenotype in prostate and colorectal cancer, as well as mitigating radiation damage to marrow. The role of EVs in leukemia and their bimodal cross talk with the encompassing microenvironment remains to be fully characterized. This may provide insight for clinical advances via the application of EVs as potential therapy and the employment of statistical and machine learning models to capture the pleiotropic effects EVs endow to a dynamic microenvironment, possibly allowing for precise therapeutic intervention. [ABSTRACT FROM AUTHOR]

Mitochondrial fragmentation frequently occurs in chronic pathological conditions as seen in various human diseases. In fact, abnormal mitochondrial morphology and mitochondrial dysfunction are hallmarks of heart failure (HF) in both human patients and HF animal models. A link between mitochondrial fragmentation and cardiac pathologies has been widely proposed, but the physiological relevance of mitochondrial fission and fusion in the heart is still unclear. Recent studies have increasingly shown that posttranslational modifications (PTMs) of fission and fusion proteins are capable of directly modulating the stability, localization, and/or activity of these proteins. These PTMs include phosphorylation, acetylation, ubiquitination, conjugation of small ubiquitin-like modifier proteins, O-linked-N-acetylglucosamine glycosylation, and proteolysis. Thus, understanding the PTMs of fission and fusion proteins may allow us to understand the complexities that determine the balance of mitochondrial fission and fusion as well as mitochondrial function in various cell types and organs including cardiomyocytes and the heart. In this review, we summarize present knowledge regarding the function and regulation of mitochondrial fission and fusion in cardiomyocytes, specifically focusing on the PTMs of each mitochondrial fission/fusion protein. We also discuss the molecular mechanisms underlying abnormal mitochondrial morphology in HF and their contributions to the development of cardiac diseases, highlighting the crucial roles of PTMs of mitochondrial fission and fusion proteins. Finally, we discuss the future potential of manipulating PTMs of fission and fusion proteins as a therapeutic strategy for preventing and/or treating HF. [ABSTRACT FROM AUTHOR]

Silicon (Si) has long been known to play a major physiological and structural role in certain organisms, including diatoms, sponges, and many higher plants, leading to the recent identification of multiple proteins responsible for Si transport in a range of algal and plant species. In mammals, despite several convincing studies suggesting that silicon is an important factor in bone development and connective tissue health, there is a critical lack of understanding about the biochemical pathways that enable Si homeostasis. Here we report the identification of a mammalian efflux Si transporter, namely Slc34a2 (also termed NaPiIIb), a known sodium-phosphate cotransporter, which was upregulated in rat kidney following chronic dietary Si deprivation. Normal rat renal epithelium demonstrated punctate expression of Slc34a2, and when the protein was heterologously expressed in Xenopus laevis oocytes, Si efflux activity (i.e., movement of Si out of cells) was induced and was quantitatively similar to that induced by the known plant Si transporter OsLsi2 in the same expression system. Interestingly, Si efflux appeared saturable over time, but it did not vary as a function of extracellular HPO42- or Na+ concentration, suggesting that Slc34a2 harbors a functionally independent transport site for Si operating in the reverse direction to the site for phosphate. Indeed, in rats with dietary Si depletion-induced upregulation of transporter expression, there was increased urinary phosphate excretion. This is the first evidence of an active Si transport protein in mammals and points towards an important role for Si in vertebrates and explains interactions between dietary phosphate and silicon. [ABSTRACT FROM AUTHOR]

Recent studies have implicated the Hippo pathway and its transcriptional effectors YAP and TAZ as necessary for fibroblast activation and tissue fibrosis. To test the specific and sufficient roles for TAZ in driving autonomous fibroblast activation, we cultured NIH3T3 fibroblasts expressing a doxycyclineinducible nuclear-localized mutant of TAZ (TAZ4SA) in scaffold-free 3D hanging drop spheroids, or on matrices of specified mechanical rigidity. Control NIH3T3 fibroblasts formed spheroids in hanging drop culture that remained stable and neither increased nor decreased in size significantly over 15 days. In contrast, TAZ4SA-transduced fibroblasts grew robustly in spheroid culture, and expressed enhanced levels of genes encoding profibrotic soluble factors connective tissue growth factor (CTGF), endothelin-1 (Et-1), and plasminogen activator inhibitor 1 (PAI-1). However, TAZ4SA expression was unable to enhance expression of extracellular matrix (ECM)-encoding genes Col1a1, Col1a2, Col3a1, or Fn1 in spheroid culture. Micromechanical testing indicated that spheroids composed of either control or TAZ4SA-expressing cells were highly compliant and indistinguishable in mechanical properties. In fibroblasts cultured on 2D matrices of compliance similar to spheroids, TAZ4SA expression was able to enhance contractile force generation, but was unable to enhance ECM gene expression. In contrast, culture on stiff hydrogels potentiated TAZ4SA enhancement of ECM expression. TAZ4SA enhancement of Col1a1 expression on soft matrices was potentiated by TGF--1, while on stiff matrices it was abrogated by inhibition of myocardin-related transcription factor, demonstrating context-dependent crosstalk of TAZ with these pathways. These findings demonstrate sufficiency of TAZ activation for driving fibroblast proliferation, contraction, and soluble profibrotic factor expression, and mechanical context-dependent crosstalk of TAZ with other pathways in regulating Col1a1 expression. [ABSTRACT FROM AUTHOR]

The mechanism responsible for the altered spectrum of tear proteins secreted by lacrimal gland acinar cells (LGAC) in patients with Sjögren's Syndrome (SS) remains unknown. We have previously identified increased cathepsin S (CTSS) activity as a unique characteristic of tears of patients with SS. Here, we investigated the role of Rab3D, Rab27a, and Rab27b proteins in the enhanced release of CTSS from LGAC. Similar to patients with SS and to the male nonobese diabetic (NOD) mouse model of SS, CTSS activity was elevated in tears of mice lacking Rab3D. Findings of lower gene expression and altered localization of Rab3D in NOD LGAC reinforce a role for Rab3D in suppressing excess CTSS release under physiological conditions. However, CTSS activity was significantly reduced in tears of mice lacking Rab27 isoforms. The reliance of CTSS secretion on Rab27 activity was supported by in vitro findings that newly synthesized CTSS was detected in and secreted from Rab27-enriched secretory vesicles and that expression of dominant negative Rab27b reduced carbachol-stimulated secretion of CTSS in cultured LGAC. High-resolution 3D-structured illumination microscopy revealed microdomains of Rab3D and Rab27 isoforms on the same secretory vesicles but present in different proportions on different vesicles, suggesting that changes in their relative association with secretory vesicles may tailor the vesicle contents. We propose that a loss of Rab3D from secretory vesicles, leading to disproportionate Rab27-to-Rab3D activity, may contribute to the enhanced release of CTSS in tears of patients with SS. [ABSTRACT FROM AUTHOR]

This study aimed to investigate protein phosphatases involved in the run down of Cav1.2 Ca2+ channels. Single ventricular myocytes obtained from adult guinea pig hearts were used to record Ca2+ channel currents with the patch-clamp technique. Calmodulin (CaM) and ATP were used to restore channel activity in inside-out patches. Inhibitors of protein phosphatases were applied to investigate the role of phosphatases. The specific protein phosphatase type 1 (PP1) inhibitor (PP1 inhibitor-2) and protein phosphatase type 2A (PP2A) inhibitor (fostriecin) abolished the slow run down of Cav1.2 Ca2+ channels, which was evident as the time-dependent attenuation of the reversing effect of CaM/ATP on the run down. However, protein phosphatase type 2B (PP2B, calcineurin) inhibitor cyclosporine A together with cyclophilin A had no effect on the channel run down. Furthermore, PP1 inhibitor-2 mainly prolonged the open time constants of the channel, specifically, the slow open time. Fostriecin primarily shortened the slow close time constants. Our data suggest that PP1 and PP2A were involved in the slow phase of Cav1.2 Ca2+ channel run down. In addition, they exerted different effects on the open-close kinetics of the channel. All above support the view that PP1 and PP2A may dephosphorylate distinct phosphorylation sites on the Cav1.2 Ca2+ channel. [ABSTRACT FROM AUTHOR]

The Na+-K+-ATPase (NKA) differs from most other ion transporters, not only in its capacity to maintain a steep electrochemical gradient across the plasma membrane, but also as a receptor for a family of cardiotonic steroids, to which ouabain belongs. Studies from many groups, performed during the last 15 years, have demonstrated that ouabain, a member of the cardiotonic steroid family, can activate a network of signaling molecules, and that NKA will also serve as a signal transducer that can provide a feedback loop between NKA and the mitochondria. This brief review summarizes the current knowledge and controversies with regard to the understanding of NKA signaling. [ABSTRACT FROM AUTHOR]

Filamin B (FLNB) is a dimeric actin-binding protein that orchestrates the reorganization of the actin cytoskeleton. Congenital mutations of FLNB at the actin-binding domain (ABD) are known to cause abnormalities of skeletal development, such as atelosteogenesis types I and III and Larsen's syndrome, although the underlying mechanisms are poorly understood. Here, using fluorescence microscopy, we characterized the reorganization of the actin cytoskeleton in cells expressing each of six pathological FLNB mutants that have been linked to skeletal abnormalities. The subfractionation assay showed a greater accumulation of the FLNB ABD mutants W148R and E227K than the wild-type protein to the cytoskeleton. Ectopic expression of FLNB-W148R and, to a lesser extent, FLNB-E227K induced prominent F-actin accumulations and the consequent rearrangement of focal adhesions, myosin II, and septin filaments and results in a delayed directional migration of the cells. The W148R protein-induced cytoskeletal rearrangement was partially attenuated by the inhibition of myosin II, p21-activated protein kinase, or Rho-associated protein kinase. The expression of a single-head ABD fragment with the mutations partially mimicked the rearrangement induced by the dimer. The F-actin clustering through the interaction with the mutant FLNB ABD may limit the cytoskeletal reorganization, preventing normal skeletal development. [ABSTRACT FROM AUTHOR]

The effectiveness and stability of epithelial barrier depend on apical junctional complexes, which consist of tight junctions (TJs) and adherens junctions (AJs). E-cadherin is the primary component of AJs, and it is essential for maintenance of cell-to-cell interactions and regulates the epithelial barrier. However, the exact mechanism underlying E-cadherin expression, particularly at the posttranscriptional level, remains largely unknown. RNA-binding proteins CUG-binding protein 1 (CUGBP1) and HU antigen R (HuR) are highly expressed in the intestinal epithelial tissues and modulate the stability and translation of target mRNAs. Here, we present evidence that CUGBP1 and HuR interact directly with the 3'-untranslated region of E-cadherin mRNA and regulate E-cadherin translation. CUGBP1 overexpression in Caco-2 cells inhibited E-cadherin translation by increasing the recruitment of E-cadherin mRNA to processing bodies (PBs), thus resulting in an increase in paracellular permeability. Overexpression of HuR exhibited an opposite effect on E-cadherin expression by preventing the translocation of E-cadherin mRNA to PBs and therefore prevented CUGBP1-induced repression of E-cadherin expression. Elevation of HuR also abolished the CUGBP1-induced epithelial barrier dysfunction. These findings indicate that CUGBP1 and HuR negate each other's effects in regulating E-cadherin translation by altering the recruitment of E-cadherin mRNA to PBs and play an important role in the regulation of intestinal barrier integrity under various pathophysiological conditions. [ABSTRACT FROM AUTHOR]

Unloading or disuse rapidly results in skeletal muscle atrophy, switching to fast-type fibers, and decreased resistance to fatigue. The recovery process is of major importance in rehabilitation for various clinical conditions. Here we studied mouse soleus muscle during 60 days of reloading after 4 wk of hindlimb suspension. Unloading produced significant atrophy of soleus muscle with decreased contractile force and fatigue resistance, accompanied by switches of myosin isoforms from IIa to IIx and IIb and fast troponin T to more low-molecular-weight splice forms. The total mass, fiber size, and contractile force of soleus muscle recovered to control levels after 15 days of reloading. However, the fatigue resistance showed a trend of worsening during this period with significant infiltration of inflammatory cells at days 3 and 7, indicating reloading injuries that were accompanied by active regeneration with upregulations of filamin-C, αB-crystallin, and desmin. The fatigue resistance partially recovered after 30-60 days of reloading. The expression of peroxisome proliferator-activated receptor γ coactivator 1α and mitofusin-2 showed changes parallel to that of fatigue resistance after unloading and during reloading, suggesting a causal role of decreased mitochondrial function. Slow fiber contents in the soleus muscle were increased after 30-60 days of reloading to become significantly higher than the normal level, indicating a secondary adaption to compensate for the slow recovery of fatigue resistance. [ABSTRACT FROM AUTHOR]

Inflammation-dominated sympathetic sprouting adjacent to the necrotic region following myocardial infarction (MI) has been implicated in the etiology of arrhythmias resulting in sudden cardiac death; however, the mechanisms responsible remain to be elucidated. Although being a key immune mediator, the role of Notch has yet to be explored. We investigated whether Notch regulates macrophage responses to inflammation and affects cardiac sympathetic reinnervation in rats undergoing MI. MI was induced by coronary artery ligation. A high level of Notch intracellular domain was observed in the macrophages that infiltrated the infarct area at 3 days post-MI. The administration of the Notch inhibitor N-N-(3,5-difluorophenacetyl-L-alanyl)-S-phenylglycine-t-butyl ester (DAPT) (intravenously 30 min before MI and then daily until death) decreased the number of macrophages and significantly increased the M2 macrophage activation profile in the early stages and attenuated the expression of nerve growth factor (NGF). Eventually, NGF-induced sympathetic hyperinnervation was blunted, as assessed by the immunofluorescence of tyrosine hydroxylase. At 7 days post-MI, the arrhythmia score of programmed electric stimulation in the vehicle-treated infarcted rats was higher than that in rats treated with DAPT. Further deterioration in cardiac function and decreases in the plasma levels of TNF-α and IL-1β were also detected. In vitro studies revealed that LPS/IFN-γ upregulated the surface expression of NGF in M1 macrophages in a Notch-dependent manner. We concluded that Notch inhibition during the acute inflammatory response phase is associated with the downregulation of NGF, probably through a macrophage-dependent pathway, thus preventing the process of sympathetic hyperinnervation. [ABSTRACT FROM AUTHOR]

Oxidative stress or reduced expression of naturally occurring antioxidants during aging has been identified as a major culprit in neuronal cell/tissue degeneration. Peroxiredoxin (Prdx) 6, a protective protein with GSH peroxidase and acidic calcium-independent phospholipase A2 activities, acts as a rheostat in regulating cellular physiology by clearing reactive oxygen species (ROS) and thereby optimizing gene regulation. We found that under stress, the neuronal cells displayed reduced expression of Prdx6 protein and mRNA with increased levels of ROS, and the cells subsequently underwent apoptosis. Using Prdx6 fused to TAT transduction domain, we showed evidence that Prdx6 was internalized in human brain cortical neuronal cells, HCN-2, and mouse hippocampal cells, HT22. The cells transduced with Prdx6 conferred resistance against the oxidative stress inducers paraquat, H2O2, and glutamate. Furthermore, Prdx6 delivery ameliorated damage to neuronal cells by optimizing ROS levels and overstimulation of NF-κB. Intriguingly, transduction of Prdx6 increased the expression of endogenous Prdx6, suggesting that protection against oxidative stress was mediated by both extrinsic and intrinsic Prdx6. The results demonstrate that Prdx6 expression is critical to protecting oxidative stress-evoked neuronal cell death. We propose that local or systemic application of Prdx6 can be an effective means of delaying/postponing neuronal degeneration. [ABSTRACT FROM AUTHOR]

The tumor microenvironment is a complex system, playing an important role in tumor development and progression. Besides cellular stromal components, extracellular matrix fibers, cytokines, and other metabolic mediators are also involved. In this review we outline the potential role of hypoxia, a major feature of most solid tumors, within the tumor microenvironment and how it contributes to immune resistance and immune suppression/tolerance and can be detrimental to antitumor effector cell functions. We also outline how hypoxic stress influences immunosuppressive pathways involving macrophages, myeloid-derived suppressor cells, T regulatory cells, and immune checkpoints and how it may confer tumor resistance. Finally, we discuss how microenvironmental hypoxia poses both obstacles and opportunities for new therapeutic immune interventions. [ABSTRACT FROM AUTHOR]

GLUT12 is a member of the facilitative family of glucose transporters. The goal of this study was to characterize the functional properties of GLUT12, expressed in Xenopus laevis oocytes, using radiotracer and electrophysiological methods. Our results showed that GLUT12 is a facilitative sugar transporter with substrate selectivity: D-glucose ≥ α-methyl-D-glucopyranoside (α-MG) > 2-deoxy-D-glucose(2-DOG) > D-fructose = D-galactose. a-MG is a characteristic substrate of the Na+/glucose (SGLT) family and has not been shown to be a substrate of any of the GLUTs. In the absence of sugar, 22Na+ was transported through GLUT12 at a higher rate (40%) than noninjected oocytes, indicating that there is a Na+ leak through GLUT12. Genistein, an inhibitor of GLUT1, also inhibited sugar uptake by GLUT12. Glucose uptake was increased by the PKA activator 8-bromoadenosine 3',5'-cyclic monophosphate (8-Br-cAMP) but not by the PKC activator phorbol-12- myristate-13-acetate (PMA). In high K+ concentrations, glucose uptake was blocked. Addition of glucose to the external solution induced an inward current with a reversal potential of approximately --15 mV and was blocked by Cl-- channel blockers, indicating the current was carried by Cl-- ions. The sugar-activated Cl-- currents were unaffected by genistein. In high external K+ concentrations, sugar-activated Cl-- currents were also blocked, indicating that GLUT12 activity is voltage dependent. Furthermore, glucose-induced current was increased by the PKA activator 8-Br-cAMP but not by the PKC activator PMA. These new features of GLUT12 are very different from those described for other GLUTs, indicating that GLUT12 must have a specific physiological role within glucose homeostasis, still to be discovered. [ABSTRACT FROM AUTHOR]

In whole cell patch-clamp recordings, we characterized the L-type Ca2+ currents in bovine adrenal zona fasciculata (AZF) cells and explored their role, along with the role of T-type channels, in ACTH- and angiotensin II (ANG II)-stimulated cortisol secretion. Two distinct dihydropyridine-sensitive L-type currents were identified, both of which were activated at relatively hyperpolarized potentials. One activated with rapid kinetics and, in conjunction with Northern blotting and PCR, was determined to be Cav1.3. The other, expressed in approximately one-half of AZF cells, activated with extremely slow voltage-dependent kinetics and combined properties not previously reported for an L-type Ca2+ channel. The T-type Ca2+ channel antagonist 3,5-dichloro-N-[1-(2,2-dimethyl-tetrahydro-pyran-4- ylmethyl)-4-fluoro-piperidin-4-ylmethyl]-benzamide (TTA-P2) inhibited Cav3.2 current in these cells, as well as ACTH- and ANG II-stimulated cortisol secretion, at concentrations that did not affect L-type currents. In contrast, nifedipine specifically inhibited L-type currents and cortisol secretion, but less effectively than TTA-P2. Diphenylbutylpiperidine Ca2+ antagonists, including pimozide, penfluridol, and fluspirilene, and the dihydropyridine niguldipine blocked Cav3.2 and L-type currents and inhibited ACTH-stimulated cortisol secretion with similar potency. This study shows that bovine AZF cells express three Ca2+ channels, the voltage-dependent gating and kinetics of which could orchestrate complex mechanisms linking peptide hormone receptors to cortisol secretion through action potentials or sustained depolarization. The function of the novel, slowly activating L-type channel is of particular interest in this respect. Regardless, the well-correlated selective inhibition of T- and L-type currents and ACTH- and ANG II-stimulated cortisol secretion by TTA-P2 and nifedipine establish the critical importance of these channels in AZF cell physiology. [ABSTRACT FROM AUTHOR]

Pituitary adenylate cyclase- activating polypeptide (PACAP) is a potent intercellular signaling molecule involved in multiple homeostatic functions. PACAP/ PAC1 receptor signaling increases excitability of neurons within the guinea pig cardiac ganglia, making them a unique system to establish mechanisms underlying PACAP modulation of neuronal function. Calcium influx is required for the PACAP-increased cardiac neuron excitability, although the pathway is unknown. This study tested whether PACAP enhancement of calcium influx through either T-type or R-type channels contributed to the modulation of excitability. Real-time quantitative polymerase chain reaction analyses indicated transcripts for Cav3.1, Cav3.2, and Cav3.3 T-type isoforms and R-type Cav2.3 in cardiac neurons. These neurons often exhibit a hyperpolarization- induced rebound depolarization that remains when cesium is present to block hyperpolarization-activated nonselective cationic currents (Ih). The T-type calcium channel inhibitors, nickel (Ni2+) or mibefradil, suppressed the rebound depolarization, and treatment with both drugs hyperpolarized cardiac neurons by 2-4 mV. Together, these results are consistent with the presence of functional T-type channels, potentially along with R-type channels, in these cardiac neurons. Fifty micromolar Ni2+, a concentration that suppresses currents in both T-type and R-type channels, blunted the PACAPinitiated increase in excitability. Ni2+ also blunted PACAP enhancement of the hyperpolarization-induced rebound depolarization and reversed the PACAP-mediated increase in excitability, after being initiated, in a subset of cells. Lastly, low voltage-activated currents, measured under perforated patch whole cell recording conditions and potentially flowing through T-type or R-type channels, were enhanced by PACAP. Together, our results suggest that a PACAP-enhanced, Ni2+-sensitive current contributes to PACAP-induced modulation of neuronal excitability. [ABSTRACT FROM AUTHOR]

We recently showed that progesterone treatment abolished arrhythmias and sudden cardiac death in a transgenic rabbit model of long QT syndrome type 2 (LQT2). Moreover, levels of cardiac sarco(endo) plasmic reticulum Ca2+-ATPase type 2a (SERCA2a) were upregulated in LQT2 heart extracts. We hypothesized that progesterone treatment upregulated SERCA2a expression, thereby reducing Ca2+-dependent arrhythmias in LQT2 rabbits. We therefore investigated the effect of progesterone on SERCA2a regulation in isolated cardiomyocytes. Cardiomyocytes from neonatal (3- to 5-day-old) rabbits were isolated, cultured, and treated with progesterone and other pharmacological agents. Immunoblotting was performed on total cell lysates and sarcoplasmic reticulum-enriched membrane fractions for protein abundance, and mRNA transcripts were quantified using real-time PCR. The effect of progesterone on baseline Ca2+ transients and Ca2+ clearance was determined using digital imaging. Progesterone treatment increased the total pool of SERCA2a protein by slowing its degradation. Using various pharmacological inhibitors of degradation pathways, we showed that progesterone-associated degradation of SERCA2a involves ubiquitination, and progesterone significantly decreases the levels of ubiquitin-tagged SERCA2a polypeptides. Our digital imaging data revealed that progesterone significantly shortened the decay and duration of Ca2+ transients. Progesterone treatment increases protein levels and activity of SERCA2a. Progesterone stabilizes SERCA2a, in part, by decreasing the ubiquitination level of SERCA2a polypeptides. [ABSTRACT FROM AUTHOR]

The endoplasmic reticulum (ER)-induced unfolded protein response (UPR) is an adaptive mechanism that is activated upon accumulation of misfolded proteins in the ER and aims at restoring ER homeostasis. In the past 10 years, the UPR has emerged as an important actor in the different phases of tumor growth. The UPR is transduced by three major ER resident stress sensors, which are protein kinase RNA-like ER kinase (PERK), activating transcription factor 6 (ATF6), and inositol- requiring enzyme-1 (IRE1). The signaling pathways elicited by those stress sensors have connections with metabolic pathways and with other plasma membrane receptor signaling networks. As such, the ER has an essential position as a signal integrator in the cell and is instrumental in the different phases of tumor progression. Herein, we describe and discuss the characteristics of an integrated signaling network that might condition the UPR biological outputs in a tissue- or stress-dependent manner. We discuss these issues in the context of the pathophysiological roles of UPR signaling in cancers. [ABSTRACT FROM AUTHOR]

The airway epithelium provides a barrier that separates inhaled air and its various particulates from the underlying tissues. It provides key physiological functions in both sensing the environment and initiating appropriate innate immune defenses to protect the lung. Protease-activated receptor-2 (PAR2) is expressed both apically and basolaterally throughout the airway epithelium. One consequence of basolateral PAR2 activation is the rapid, Ca2+-dependent ion flux that favors secretion in the normally absorptive airway epithelium. However, roles for apically expressed PAR2 activation have not been demonstrated, in part due to the lack of specific, high-potency PAR2 ligands. In the present study, we used the newly developed PAR2 ligand 2at-LIGRLO(PEG3-Pam)-NH2 in combination with well-differentiated, primary cultured airway epithelial cells from wild-type and PAR2-/- mice to examine the physiological role of PAR2 in the conducting airway after apical activation. Using digital imaging microscopy of intracellular Ca2+ concentration changes, we verified ligand potency on PAR2 in primary cultured airway cells. Examination of airway epithelial tissue in an Ussing chamber showed that apical activation of PAR2 by 2at-LIGRLO(PEG3-Pam)-NH2 resulted in a transient decrease in transepithelial resistance that was due to increased apical ion efflux. We determined pharmacologically that this increase in ion conductance was through Ca2+-activated Cl- and large-conductance K+ channels that were blocked with a Ca2+- activated Cl- channel inhibitor and clotrimazole, respectively. Stimulation of Cl- efflux via PAR2 activation at the airway epithelial surface can increase airway surface liquid that would aid in clearing the airway of noxious inhaled agents. [ABSTRACT FROM AUTHOR]

A reversible inhibition of mitochondrial respiration by complex I inhibition at the onset of reperfusion decreases injury in buffer-perfused hearts. Administration of acidic reperfusate for a brief period at reperfusion decreases cardiac injury. We asked if acidification treatment decreased cardiac injury during reperfusion by inhibiting complex I. Exposure of isolated mouse heart mitochondria to acidic buffer decreased the complex I substrate-stimulated respiration, whereas respiration with complex II substrates was unaltered. Evidence of the rapid and reversible inhibition of complex I by an acidic environment was obtained at the level of isolated complex, intact mitochondria and in situ mitochondria in digitonin-permeabilized cardiac myocytes. Moreover, ischemia-damaged complex I was also reversibly inhibited by an acidic environment. In the buffer-perfused mouse heart, reperfusion with pH 6.6 buffer for the initial 5 min decreased infarction. Compared with untreated hearts, acidification treatment markedly decreased the mitochondrial generation of reactive oxygen species and improved mitochondrial calcium retention capacity and inner mitochondrial membrane integrity. The decrease in infarct size achieved by acidic reperfusion approximates the reduction obtained by a reversible, partial blockade of complex I at reperfusion. Extracellular acidification decreases cardiac injury during reperfusion in part via the transient and reversible inhibition of complex I, leading to a reduction of oxyradical generation accompanied by a decreased susceptibility to mitochondrial permeability transition during early reperfusion. [ABSTRACT FROM AUTHOR]

Due to evolutionary pressure, there are many complex interactions at the interface between pathogens and eukaryotic host cells wherein host cells attempt to clear invading microorganisms and pathogens counter these mechanisms to colonize and invade host tissues. One striking observation from studies focused on this interface is that pathogens have multiple mechanisms to modulate and disrupt normal cellular physiology to establish replication niches and avoid clearance. The precision by which pathogens exert their effects on host cells makes them excellent tools to answer questions about cell physiology of eukaryotic cells. Furthermore, an understanding of these mechanisms at the host-pathogen interface will benefit our understanding of how pathogens cause disease. In this review, we describe a few examples of how pathogens disrupt normal cellular physiology and protein trafficking at epithelial cell barriers to underscore how pathogens modulate cellular processes to cause disease and how this knowledge has been utilized to learn about cellular physiology. [ABSTRACT FROM AUTHOR]

The formation of the first epithelium was an essential step for animal evolution, since it has allowed coordination of the behavior of a cell layer and creation of a selective barrier between the internal medium and the outside world. The possibility of coupling the cells in a single layer has allowed morphogenetic events, such as tube formation, or gastrulation, to form more complex animal morphologies. The invention of sealed junctions between cells has allowed, on the other hand, creation of an asymmetry of nutrients or salts between the apical and the basal side of the epithelial layer. Creation of an internal medium has led to homeostasis, allowing the evolution of more complex physiological functions and the emergence of sophisticated animal shapes. During evolution, the origins of the first animals coincided with the invention of several protein complexes, including true cadherins and the polarity protein complexes. How these complexes regulate formation of the apicolateral border and the adherens junctions is still not fully understood. This review focuses on the role of these apical polarity complexes and, in particular, the Crumbs complex, which is essential for proper organization of epithelial layers from Drosophila to humans [ABSTRACT FROM AUTHOR]

Indeed, cellular heterogeneity and evolution are at the core of cancer biology, as cancer cell diversity provides the fuel for tumor evolution and progression to metastasis. EDITORIAL Dynamic Tumor Heterogeneity and Cancer Progression AmericanJournalofPhysiology-CellPhysiologybeginsaThemeofReviewson DynamicTumorHeterogeneityandCancerProgression Thordur Oskarsson Heidelberg Institute for Stem Cell Technology and Experimental Medicine (HI-STEM gGmbH) and Division of Stem Cells and Cancer, German Cancer Research Center (DKFZ), Heidelberg, Germany Cancer is a heterogeneous collection of diseases that affectmostregeneratingtissues. [Extracted from the article]

Platelet-derived growth factor (PDGF) stimulates vascular smooth muscle cell (VSMC) migration and neointimal formation in response to injury. We previously identified IQ-domain GTPase-activating protein 1 (IQGAP1) as a novel VEGF receptor 2 binding scaffold protein involved in endothelial migration. However, its role in VSMC migration and neointimal formation in vivo is unknown. Here we show that PDGF stimulation rapidly promotes IQGAP1 association with PDGF receptor (PDGFR) as well as IQGAP1 tyrosine phosphorylation in cultured VSMC. Overexpression or knockdown of IQGAP1 enhances or inhibits PDGFR autophosphorylation (p-PDGFR), respectively. Immunofluorescence and cell fractionation analysis reveals that PDGF-induced p-PDGFR localized in focal adhesions (FAs), but not caveolae/lipid rafts, is inhibited by IQGAP1 knockdown with siRNA. PDGF stimulation promotes IQGAP1 association with PDGFR/FA signaling protein complex. Functionally, IQGAP1 siRNA inhibits PDGF-induced FA formation as well as VSMC migration induced by PDGF. In vivo, IQGAP1 expression is markedly increased at neointimal VSMC in wire-injured femoral arteries. Mice lacking IQGAP1 exhibit impaired neointimal formation in response to vascular injury. In summary, IQGAP1, through interaction with PDGFR and FA signaling proteins, promotes activation of PDGFR in FAs as well as FA formation, which may contribute to VSMC migration and neointimal formation after injury. Our findings provide insight into IQGAP1 as a potential therapeutic target for vascular migration-related diseases. [ABSTRACT FROM AUTHOR]

Evidence from the literature suggests that testosterone plays an important role in visceral fat accumulation since both men and women with hyperandrogenism accumulate more adipose tissue in the abdominal cavity than healthy women. However, the underlying mechanisms remain to be elucidated. To shed light on this issue, we have used an in vitro approach to examine the effect of testosterone on human visceral preadipocyte proliferation. Our results showed that testosterone treatment significantly increased proliferation of human visceral preadipocytes in proliferation assays using flow cytometric analysis. We next performed a microarray gene expression analysis of human visceral preadipocytes treated with testosterone or vehicle to identify which genes were involved in the testosterone-induced increase in preadipocyte proliferation. The results showed a total of 140 genes differentially expressed between testosterone vs. vehicle. Among the top 10 upregulated genes, 5 were involved in cellular cycle and proliferation, and 3 (APOBEC3b, CCNA2, and PRC1) were significantly overexpressed by testosterone treatment when analyzed by real-time PCR. We conclude that testosterone exerts a proliferative effect on preadipocytes that may participate in the sex differences in fat distribution and that it may explain visceral fat accumulation in women with hyperandrogenism. [ABSTRACT FROM AUTHOR]

TMEM16 (transmembrane protein 16) proteins, which possess eight putative transmembrane domains with intracellular NH2- and COOH-terminal tails, are thought to comprise a Cl- channel family. The function of TMEM16F, a member of the TMEM16 family, has been greatly controversial. In the present study, we performed whole cell patchclamp recordings to investigate the function of human TMEM16F. In TMEM16F-transfected HEK293T cells but not TMEM16K- and mock-transfected cells, activation of membrane currents with strong outward rectification was found to be induced by application of a Ca2+ ionophore, ionomycin, or by an increase in the intracellular free Ca2+ concentration. The free Ca2+ concentration for half-maximal activation of TMEM16F currents was 9.6 μM, which is distinctly higher than that for TMEM16A/B currents. The outwardly rectifying current-voltage relationship for TMEM16F currents was not changed by an increase in the intracellular Ca2+ level, in contrast to TMEM16A/B currents. The Ca2+-activated TMEM16F currents were anion selective, because replacing Cl- with aspartate- in the bathing solution without changing cation concentrations caused a positive shift of the reversal potential. The anion selectivity sequence of the TMEM16F channel was I- > Br- > Cl- > F- > aspartate-. Niflumic acid, a Ca2+-activated Cl- channel blocker, inhibited the TMEM16F-dependent Cl- currents. Neither overexpression nor knockdown of TMEM16F affected volume-sensitive outwardly rectifying Cl- channel (VSOR) currents activated by osmotic swelling or apoptotic stimulation. These results demonstrate that human TMEM16F is an essential component of a Ca2+-activated Cl- channel with a Ca2+ sensitivity that is distinct from that of TMEM16A/B and that it is not related to VSOR activity. [ABSTRACT FROM AUTHOR]

Among the over 300 members of the solute carrier (SLC) group of integral plasma membrane transport proteins are the nine electroneutral cation-chloride cotransporters belonging to the SLC12 gene family. Seven of these transporters have been functionally described as coupling the electrically silent movement of chloride with sodium and/or potassium. Although in silico analysis has identified two additional SLC12 family members, no physiological role has been ascribed to the proteins encoded by either the SLC12A8 or the SLC12A9 genes. Evolutionary conservation of this gene family from protists to humans confirms their importance. A wealth of physiological, immunohistochemical, and biochemical studies have revealed a great deal of information regarding the importance of this gene family to human health and disease. The sequencing of the human genome has provided investigators with the capability to link several human diseases with mutations in the genes encoding these plasma membrane proteins. The availability of bacterial artificial chromosomes, recombination engineering techniques, and the mouse genome sequence has simplified the creation of targeting constructs to manipulate the expression/function of these cation-chloride cotransporters in the mouse in an attempt to recapitulate some of these human pathologies. This review will summarize the three human disorders that have been linked to the mutation/dysfunction of the Na-Cl, Na-K-2Cl, and K-Cl cotransporters (i.e., Bartter's, Gitleman's, and Andermann's syndromes), examine some additional pathologies arising from genetically modified mouse models of these cotransporters including deafness, blood pressure, hyperexcitability, and epithelial transport deficit phenotypes. [ABSTRACT FROM AUTHOR]

Mast cells are one of the major producers of prostaglandins (PGs). The final metabolite of PGs, 15-deoxy-delta-12,14-PGJ2 (15-deoxy-delta PGJ2) is the endogenous ligand of the peroxisome proliferator-activated receptor (PPAR) γ. PPARγ modulates adipocyte differentiation; therefore, we attempted to investigate whether PGs derived from mast cells influenced on adipogenesis. We found the increase of mast cell numbers in fat tissue of obese mice fed with a high fat diet, allowing us to speculate contributions of mast cells to adipogenesis. Mast cell-mediated induction of adipogenesis was evaluated by using 3T3 L1 cells. Supernatants obtained from mast cells stimulated with calcium ionophore or the high glucose condition contained 15-deoxy-delta PGJ2 and induced adipogenesis of 3T3 L1 cells. Agonistic activity of PGJ2 from the supernatants on PPARγ was confirmed by a reporter gene assay. Culture medium collected from calcium ionophore-stimulated BMCMC activated PPAR responsive element in NIH3T3 fibroblasts, and the specific inhibitor of PPARγ canceled the activation. Contribution of mast cells to obesity was evaluated by using mast cell-deficient mice fed with a western diet. Weight gain of mast cell-deficient mice during high-fat feeding was impaired as compared to their littermate wild type mice; on the other hand, transplantation of bone marrow-derived cultured mast cells to mast cell-deficient mice restored the weight gain by intake of a high-fat diet. In this study, we clearly demonstrated that mast cells produced PGs in response to the high glucose condition and induced adipocyte differentiation and possibly obesity. This is the first study that provides evidence for a novel role of mast cells in adipogenesis via PPARγ activation. [ABSTRACT FROM AUTHOR]

TM4SF10 is a claudin-like cell junction protein that is transiently expressed during podocyte development where its expression is downregulated in differentiating podocytes coincident with the appearance of Nephrin at the slit diaphragm. In a yeast two hybrid screen, we identified ADAP, a well known Fyn substrate and Fyn binding partner, as a TM4SF10 interacting protein in mouse kidney. Using co-immunoprecipitation and immunohistochemistry experiments in cultured human podocytes, we show that TM4SF10 co-localizes with Fyn and ADAP, but does not form a stable complex with Fyn. Cytoskeletal changes and phosphorylation events mediated by Fyn activity were reversed by TM4SF10 over-expression, including a decrease in the activating tyrosine phosphorylation of Fyn (Y421) suggesting TM4SF10 may have a regulatory role in suppressing Fyn activity. In addition, TM4SF10 expression enhanced the abundance of high molecular weight forms of Nephrin indicating it may participate in a mechanism controlling Nephrin's appearance at the plasma membrane. Therefore, these studies have identified ADAP as another Fyn adapter protein expressed in podocytes, and that TM4SF10, possibly through ADAP, may regulate Fyn activity. Since TM4SF10 expression is temporally regulated during kidney development, these studies may help define a mechanism by which the slit diaphragm matures as a highly specialized cell junction during podocyte differentiation. [ABSTRACT FROM AUTHOR]

Overactivation of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels is involved in diabetes-depressed excitability of aortic baroreceptor neurons in nodose ganglia. This involvement links to the autonomic dysfunction associated with high morbidity and mortality in diabetic patients. The present study examined the effects of an angiotensin II type I receptor (AT1R) antagonist (losartan), a NADPH oxidase inhibitor (apocynin), and a superoxide dismutase mimetic (tempol) on the enhanced HCN currents and attenuated cell excitability in diabetic nodose neurons. In sham and streptozotocin-induced type 1 diabetic rats, HCN currents and cell excitability of aortic baroreceptor neurons were recorded by the whole cell patch-clamp technique. The angiotensin II level in nodose ganglia from diabetic rats was higher than that from sham rats (101.6 ± 4.8 vs. 38.9 ± 4.2 pg/mg protein, p<0.05). Single-cell RT-PCR, western blot, immunofluorescence staining, and chemiluminescence data showed that mRNA and protein expression of AT1R, protein expression of NADPH oxidase components, and superoxide production in nodose neurons were increased in diabetic rats as compared with those from sham rats. HCN current density was higher and cell excitability was lower in aortic baroreceptor neurons from diabetic rats than that from sham rats. Losartan (1 μM), apocynin (100 μM), and tempol (1 mM) normalized the enhanced HCN current density and increased the cell excitability in the aortic baroreceptor neurons of diabetic rats. These findings suggest that endogenous angiotensin II-NADPH oxidase-superoxide signaling contributes to the enhanced HCN currents and the depressed cell excitation in the aortic baroreceptor neurons of diabetic rats. [ABSTRACT FROM AUTHOR]