Showing papers in "The FASEB Journal in 1999"
TL;DR: Recent developments that have widened considerably the understanding of the mechanisms that control V EGF production and VEGF signal transduction are focused on and recent studies that have shed light on the mechanisms by which VEGf regulates angiogenesis are reviewed.
Abstract: Vascular endothelial growth factor (VEGF) is a highly specific mitogen for vascular endothelial cells. Five VEGF isoforms are generated as a result of alternative splicing from a single VEGF gene. These isoforms differ in their molecular mass and in biological properties such as their ability to bind to cell-surface heparan-sulfate proteoglycans. The expression of VEGF is potentiated in response to hypoxia, by activated oncogenes, and by a variety of cytokines. VEGF induces endothelial cell proliferation, promotes cell migration, and inhibits apoptosis. In vivo VEGF induces angiogenesis as well as permeabilization of blood vessels, and plays a central role in the regulation of vasculogenesis. Deregulated VEGF expression contributes to the development of solid tumors by promoting tumor angiogenesis and to the etiology of several additional diseases that are characterized by abnormal angiogenesis. Consequently, inhibition of VEGF signaling abrogates the development of a wide variety of tumors. The various VEGF forms bind to two tyrosine-kinase receptors, VEGFR-1 (flt-1) and VEGFR-2 (KDR/flk-1), which are expressed almost exclusively in endothelial cells. Endothelial cells express in addition the neuropilin-1 and neuropilin-2 coreceptors, which bind selectively to the 165 amino acid form of VEGF (VEGF165). This review focuses on recent developments that have widened considerably our understanding of the mechanisms that control VEGF production and VEGF signal transduction and on recent studies that have shed light on the mechanisms by which VEGF regulates angiogenesis.
3,569 citations
TL;DR: Current knowledge on the function of vitamin E is summarized, with emphasis on its antioxidant vs. other properties, the preference of the organism for RRR‐α‐tocopherol, and its metabolism to CEHCs.
Abstract: Although vitamin E has been known as an essential nutrient for reproduction since 1922, we are far from understanding the mechanisms of its physiological functions. Vitamin E is the term for a group of tocopherols and tocotrienols, of which alpha-tocopherol has the highest biological activity. Due to the potent antioxidant properties of tocopherols, the impact of alpha-tocopherol in the prevention of chronic diseases believed to be associated with oxidative stress has often been studied, and beneficial effects have been demonstrated. Recent observations that the alpha-tocopherol transfer protein in the liver specifically sorts out RRR-alpha-tocopherol from all incoming tocopherols for incorporation into plasma lipoproteins, and that alpha-tocopherol has signaling functions in vascular smooth muscle cells that cannot be exerted by other forms of tocopherol with similar antioxidative properties, have raised interest in the roles of vitamin E beyond its antioxidative function. Also, gamma-tocopherol might have functions apart from being an antioxidant. It is a nucleophile able to trap electrophilic mutagens in lipophilic compartments and generates a metabolite that facilitates natriuresis. The metabolism of vitamin E is equally unclear. Excess alpha-tocopherol is converted into alpha-CEHC and excreted in the urine. Other tocopherols, like gamma- and delta-tocopherol, are almost quantitatively degraded and excreted in the urine as the corresponding CEHCs. All rac alpha-tocopherol compared to RRR-alpha-tocopherol is preferentially degraded to alpha-CEHC. Thus, there must be a specific, molecular role of RRR-alpha-tocopherol that is regulated by a system that sorts, distributes, and degrades the different forms of vitamin E, but has not yet been identified. In this article we try to summarize current knowledge on the function of vitamin E, with emphasis on its antioxidant vs. other properties, the preference of the organism for RRR-alpha-tocopherol, and its metabolism to CEHCs.
1,516 citations
TL;DR: The role of mitogen‐activated protein kinases and AP‐1 and ETS transcription factors in the regulation of MMP gene expression during invasion process is focused on.
Abstract: Degradation of basement membranes and stromal extracellular matrix (ECM) is crucial for invasion and metastasis of malignant cells. Degradation of ECM is initiated by proteinases secreted by different cell types participating in tumor cell invasion, and increased expression or activity of every known class of proteinases (metallo-, serine-, aspartic-, and cysteine) has been linked to malignancy and invasion of tumor cells. Studies performed over the last decade have revealed that matrix metalloproteinases (MMPs) play a crucial role in tumor invasion. Expression of MMP genes is transcriptionally regulated by a variety of extracellular factors including cytokines, growth factors, and cell contact to ECM. This review will summarize the current view on the role of MMPs in tumor growth, invasion, and survival, and focus on the role of mitogen-activated protein kinases and AP-1 and ETS transcription factors in the regulation of MMP gene expression during invasion process.
1,508 citations
TL;DR: Although the precise mechanisms by which nonparenchymal cells modulate the hepatocyte phenotype remain unelucidated, some new insights on the modes of cell signaling, the extent of cell–cell interaction, and the ratio of cell populations are noted.
Abstract: Heterotypic cell interaction between parenchymal cells and nonparenchymal neighbors has been reported to modulate cell growth, migration, and/or differentiation. In both the developing and adult liver, cell-cell interactions are imperative for coordinated organ function. In vitro, cocultivation of hepatocytes and nonparenchymal cells has been used to preserve and modulate the hepatocyte phenotype. We summarize previous studies in this area as well as recent advances in microfabrication that have allowed for more precise control over cell-cell interactions through 'cellular patterning' or 'micropatterning'. Although the precise mechanisms by which nonparenchymal cells modulate the hepatocyte phenotype remain unelucidated, some new insights on the modes of cell signaling, the extent of cell-cell interaction, and the ratio of cell populations are noted. Proposed clinical applications of hepatocyte cocultures, typically extracorporeal bioartificial liver support systems, are reviewed in the context of these new findings. Continued advances in microfabrication and cell culture will allow further study of the role of cell communication in physiological and pathophysiological processes as well as in the development of functional tissue constructs for medical applications.
903 citations
TL;DR: Burger et al. as mentioned in this paper showed that the role of osteocytes as professional mechanosensory cells of bone, and the lacuno-canalicular porosity as the structure that mediates mechanosensing, can explain local bone gain and loss, as well as remodeling in response to fatigue damage.
Abstract: The capacity of bone tissue to alter its mass and structure in response to mechanical demands has long been recognized but the cellular mechanisms involved remained poorly understood. Over the last several years significant progress has been made in this field, which we will try to summarize. These studies emphasize the role of osteocytes as the professional mechanosensory cells of bone, and the lacuno-canalicular porosity as the structure that mediates mechanosensing. Strain-derived flow of interstitial fluid through this porosity seems to mechanically activate the osteocytes, as well as ensuring transport of cell signaling molecules and nutrients and waste products. This concept allows an explanation of local bone gain and loss, as well as remodeling in response to fatigue damage, as processes supervised by mechanosensitive osteocytes.—Burger, E. H., Klein-Nulend, J. Mechanotransduction in bone—role of the lacuno-canalicular network.
901 citations
TL;DR: This work has shown that both transcriptional and post‐transcriptional mechanisms modulate the activity of this critical cellular enzyme, glutathione, in a tightly regulated manner in mammalian cells.
Abstract: Glutathione (GSH) is an important intracellular peptide with multiple functions ranging from antioxidant defense to modulation of cell proliferation. GSH is synthesized in the cytosol of all mammalian cells in a tightly regulated manner. The major determinants of GSH synthesis are the avail- ability of cysteine, the sulfur amino acid precursor, and the activity of the rate-limiting enzyme, g-glu- tamylcysteine synthetase (GCS). In the liver, major factors that determine the availability of cysteine are diet, membrane transport activities of the three sulfur amino acids cysteine, cystine and methionine, and the conversion of methionine to cysteine via the trans-sulfuration pathway. Many conditions alter GSH level via changes in GCS activity and GCS gene expression. These include oxidative stress, activators of Phase II detoxifying enzymes, antioxidants, drug- resistant tumor cell lines, hormones, cell prolifera- tion, and diabetes mellitus. Since the molecular cloning of GCS, much has been learned about the regulation of this enzyme. Both transcriptional and post-transcriptional mechanisms modulate the activ- ity of this critical cellular enzyme.—Lu, S. C. Regu- lation of hepatic glutathione synthesis: current con- cepts and controversies. FASEB J. 13, 1169 -1183 (1999)
862 citations
TL;DR: Although the data on protein oxidation in humans are sparse and inconclusive, the available data in animals consistently show an antioxidant role of vitamin C, both in the presence and absence of metal ions.
Abstract: Vitamin C readily scavenges reactive oxygen and nitrogen species and may thereby prevent oxidative damage to important biological macromolecules such as DNA, lipids, and proteins. Vitamin C also reduces redox active transition metal ions in the active sites of specific biosynthetic enzymes. The interaction of vitamin C with ‘free’, catalytically active metal ions could contribute to oxidative damage through the production of hydroxyl and alkoxyl radicals; whether these mechanisms occur in vivo, however, is uncertain. To examine this issue, we reviewed studies that investigated the role of vitamin C, both in the presence and absence of metal ions, in oxidative DNA, lipid, and protein damage. We found compelling evidence for antioxidant protection of lipids by vitamin C in biological fluids, animals, and humans, both with and without iron cosupplementation. Although the data on protein oxidation in humans are sparse and inconclusive, the available data in animals consistently show an antioxidant role of vit...
836 citations
TL;DR: This work will focus on recent insights into the molecular mechanisms by which signaling specificity between different TGF‐β superfamily members is achieved and regulated, and how a single family member can elicit a broad scala of biological responses.
Abstract: Transforming growth factor-β (TGF-β) superfamily members are multifunctional cell–cell signaling proteins that play pivotal roles in tissue homeostasis and development of multicellular animals. They mediate their pleiotropic effects from membrane to nucleus through distinct combinations of type I and type II serine/threonine kinase receptors and their downstream effectors, known as Smad proteins. Certain Smads, termed receptor-regulated Smads, become phosphorylated by activated type I receptors and form heteromeric complexes with a common-partner Smad4, which translocates into the nucleus to control gene transcription. In addition to these signal transducing Smads, inhibitory Smads have been identified that inhibit the activation of receptor-regulated Smads. In contrast to the still growing TGF-β superfamily (with ∼30 members in mammals), relatively few type I and type II receptors as well as Smads have been identified. We will focus on recent insights into the molecular mechanisms by which signaling spec...
806 citations
TL;DR: Recent studies have advanced the understanding of the signal transduction pathway leading to NF‐κB activation by cytokines and will provide insights for the mechanism by which NF-κB is regulated by oxidative stress.
Abstract: NF-kappaB is a dimeric transcription factor that is involved in the regulation of a large number of genes that control various aspects of the immune and inflammatory response. It is activated by a variety of stimuli ranging from cytokines, to various forms of radiation, to oxidative stress (such as exposure to H2O2). Recent studies have advanced our understanding of the signal transduction pathway leading to NF-kappaB activation by cytokines and will provide insights for the mechanism by which NF-kappaB is regulated by oxidative stress. An important question that is yet to be answered is whether reactive oxygen species play a physiological role in NF-kappaB activation.
796 citations
TL;DR: It is found that the mRNA level of the sod‐3 gene, which encodes Mn‐superoxide dismutase (SOD), was much higher in daf‐2 mutants than in the wild type, suggesting that the daf-2 gene network controls longevity by regulating the Mn‐SOD‐associated antioxidant defense system.
Abstract: Longevity is regulated by the daf-2 gene network in Caenorhabditis elegans. Mutations in the daf-2 gene, which encodes a member of the insulin receptor family, confer the life extension (Age) phenotype and the constitutive dauer (a growth-arrested larval form specialized for dispersal) formation phenotype. The Age phenotype is mutually potentiated by two life extension mutations in the daf-2 gene and the clk-1 gene, a homologue of yeast CAT5/COQ7 known to regulate ubiquinone biosynthesis. In this study, we demonstrated that the daf-2 mutation also conferred an oxidative stress resistance (Oxr) phenotype, which was also enhanced by the clk-1 mutation. Similar to the Age phenotype, the Oxr phenotype was regulated by the genetic pathway of insulin-like signaling from daf-2 to the daf-16 gene, a homologue of the HNF-3/forkhead transcription factor. These findings led us to examine whether the insulin-like signaling pathway regulates the gene expression of antioxidant defense enzymes. We found that the mRNA le...
747 citations
TL;DR: It is demonstrated that ROS are second messengers involved in leptin‐induced signaling in endothelial cells, which may activate atherogenic processes and contribute to the development of vascular pathology.
Abstract: Human umbilical vein endothelial cells (HUVEC) express functional receptors to leptin, the product of the ob gene. As human obesity is associated with atherosclerosis and hyperleptinemia, we investigated whether leptin, in addition to its angiogenic properties, exerts atherogenic effects through the generation of oxidative stress in endothelial cells. In HUVEC leptin increased the accumulation of reactive oxygen species (ROS), as assessed by the oxidation of 2', 7'- dichlorodihydrofluorescein, in a time- and concentration-dependent manner. In addition, leptin activated the NH2-terminal c-Jun kinase/stress-activated protein kinase pathway as demonstrated by enhanced JNK activity and AP-1 DNA binding. Both effects were sensitive to antioxidant treatment with N-acetylcysteine. NF-kappaB, another redox-sensitive transcription factor, was also activated by leptin stimulation in an oxidant-dependent manner. Finally, activation of both AP-1 and NF-kappaB was associated with an enhanced expression of the monocyte chemoattractant protein-1 in HUVEC. These findings demonstrate that ROS are second messengers involved in leptin-induced signaling in endothelial cells. Thus, chronic oxidative stress in endothelial cells under hyperleptinemia may activate atherogenic processes and contribute to the development of vascular pathology.
TL;DR: In vitro and in vivo studies indicate that CTGF mediates TGF‐β‐induced fibroblast collagen synthesis and that in vivo blockade of CTGF synthesis or action reduces TGF-β‐ induced granulation tissue formation by inhibiting both collagen synthesisation and fibro Blast accumulation.
Abstract: Connective tissue growth factor (CTGF) is a cysteine-rich peptide synthesized and secreted by fibroblastic cells after activation with transforming growth factor beta (TGF-beta) that acts as a downstream mediator of TGF-beta-induced fibroblast proliferation. We performed in vitro and in vivo studies to determine whether CTGF is also essential for TGF-beta-induced fibroblast collagen synthesis. In vitro studies with normal rat kidney (NRK) fibroblasts demonstrated CTGF potently induces collagen synthesis and transfection with an antisense CTGF gene blocked TGF-beta stimulated collagen synthesis. Moreover, TGF-beta-induced collagen synthesis in both NRK and human foreskin fibroblasts was effectively blocked with specific anti-CTGF antibodies and by suppressing TGF-beta-induced CTGF gene expression by elevating intracellular cAMP levels with either membrane-permeable 8-Br-cAMP or an adenylyl cyclase activator, cholera toxin (CTX). cAMP also inhibited collagen synthesis induced by CTGF itself, in contrast to its previously reported lack of effect on CTGF-induced DNA synthesis. In animal assays, CTX injected intradermally in transgenic mice suppressed TGF-beta activation of a human CTGF promoter/lacZ reporter transgene. Both 8-Br-cAMP and CTX blocked TGF-beta-induced collagen deposition in a wound chamber model of fibrosis in rats. CTX also reduced dermal granulation tissue fibroblast population increases induced by TGF-beta in neonatal mice, but not increases induced by CTGF or TGF-beta combined with CTGF. Our data indicate that CTGF mediates TGF-beta-induced fibroblast collagen synthesis and that in vivo blockade of CTGF synthesis or action reduces TGF-beta-induced granulation tissue formation by inhibiting both collagen synthesis and fibroblast accumulation.
TL;DR: It is speculated that some of the biological responses elicited by phorbol esters or by activation of receptors coupled to elevation in DAG levels could be mediated by PKC‐independent pathways.
Abstract: Protein kinase C (PKC), a family of related serine-threonine kinases, is a key player in the cellular responses mediated by the second messenger diacylglycerol (DAG) and the phorbol ester tumor promoters. The traditional view of PKCs as DAG/phospholipid-regulated proteins has expanded in the last few years by three seminal discoveries. First, PKC activity and maturation is controlled by autophosphorylation and transphosphorylation mechanisms, which includes phosphorylation of PKC isozymes by phosphoinositide-dependent protein kinases (PDKs) and tyrosine kinases. Second, PKC activity and localization are regulated by direct interaction with different types of interacting proteins. Protein-protein interactions are now recognized as important mechanisms that target individual PKCs to different intracellular compartments and confer selectivity by associating individual isozymes with specific substrates. Last, the discovery of novel phorbol ester receptors lacking kinase activity allows us to speculate that some of the biological responses elicited by phorbol esters or by activation of receptors coupled to elevation in DAG levels could be mediated by PKC-independent pathways.
TL;DR: Using transgenic mice that overexpress aldose reductase in their lenses, it is found that the flux of glucose through the polyol pathway is the major cause of hyperglycemic oxidative stress in this tissue.
Abstract: There is strong evidence to show that diabetes is associated with increased oxidative stress. However, the source of this oxidative stress remains unclear. Using transgenic mice that overexpress aldose reductase (AR) in their lenses, we found that the flux of glucose through the polyol pathway is the major cause of hyperglycemic oxidative stress in this tissue. The substantial decrease in the level of reduced glutathione (GSH) with concomitant rise in the level of lipid peroxidation product malondialdehyde (MDA) in the lens of transgenic mice, but not in the nontransgenic mice, suggests that glucose autoxidation and nonenzymatic glycation do not contribute significantly to oxidative stress in diabetic lenses. AR reduction of glucose to sorbitol probably contributes to oxidative stress by depleting its cofactor NADPH, which is also required for the regeneration of GSH. Sorbitol dehydrogenase, the second enzyme in the polyol pathway that converts sorbitol to fructose, also contributes to oxidative stress, m...
TL;DR: It is demonstrated that HSP27 overexpression decreases U937 human leukemic cell sensitivity to etoposide‐induced cytotoxicity by preventing apoptosis, and also prevents procaspase‐9 activation.
Abstract: We have previously shown that the small heat shock protein HSP27 inhibited apoptotic pathways triggered by a variety of stimuli in mammalian cells. The present study demonstrates that HSP27 overexpression decreases U937 human leukemic cell sensitivity to etoposide-induced cytotoxicity by preventing apoptosis. As observed for Bcl-2, HSP27 overexpression delays poly(ADP-ribose)polymerase cleavage and procaspase-3 activation. In contrast with Bcl-2, HSP27 overexpression does not prevent etoposide-induced cytochrome c release from the mitochondria. In a cell-free system, addition of cytochrome c and dATP to cytosolic extracts from untreated cells induces the proteolytic activation of procaspase-3 in both control and bcl-2-transfected U937 cells but fails to activate procaspase-3 in HSP27-overexpressing cells. Immunodepletion of HSP27 from cytosolic extracts increases cytochrome c/dATP-mediated activation of procaspase-3. Overexpression of HSP27 also prevents procaspase-9 activation. In the cell-free system, immunodepletion of HSP27 increases LEDH-AFC peptide cleavage activity triggered by cytochrome c/dATP treatment. We conclude that HSP27 inhibits etoposide-induced apoptosis by preventing cytochrome c and dATP-triggered activity of caspase-9, downstream of cytochrome c release.
TL;DR: Nucleolin is a major nucleolar protein of exponentially growing eukaryotic cells, which is directly involved in the regulation of ribosome biogenesis and maturation, and is a target for regulation by proteolysis, methylation, ADP‐ribosylation, and phosphorylation by CKII, cdc2, PKC‐ξ, cyclic AMP‐dependent protein kinase, and ecto‐protein kinase.
Abstract: Cells require optimum protein synthetic activity in order to support cell proliferation, maintain homeostatic and metabolic integrity, and repair damage. Since growth depends on protein synthesis through ribosome biogenesis, the control of biosynthesis of ribosomes is necessarily a key element for control of growth. Nucleolin is a major nucleolar protein of exponentially growing eukaryotic cells, which is directly involved in the regulation of ribosome biogenesis and maturation. The highly conserved nucleolin contains three major domains through which it controls the organization of nucleolar chromatin, packaging of pre-RNA, rDNA transcription, and ribosome assembly. Numerous reports have implicated the involvement of nucleolin either directly or indirectly in the regulation of cell proliferation and growth, cytokinesis, replication, embryogenesis, and nucleogenesis. Nucleolin, an RNA binding protein, is also an autoantigen, a transcriptional repressor, and a switch region targeting factor. In addition, n...
TL;DR: It is demonstrated that SDF‐1α activation promotes the physical association of Gαi with the CXCR4, and compelling evidence is presented that the chemokines signal through mechanisms similar to those activated by cytokines.
Abstract: The chemokine stromal cell-derived factor (SDF-1α), the ligand for the CXCR4 receptor, induces a wide variety of effects that include calcium mobilization, chemotactic responses, bone marrow myelopoiesis, neuronal patterning, and prevention of HIV-1 infection. Nonetheless, little is known of the biochemical pathways required to achieve this variety of responses triggered after receptor–chemokine interaction. We developed a set of monoclonal antibodies that specifically recognize the CXCR4 receptor and used them to identify the signaling pathway activated after SDF-1α binding in human T cell lines. Here we demonstrate that SDF-1α activation promotes the physical association of Gαi with the CXCR4. Furthermore, within seconds of SDF-1α activation, the CXCR4 receptor becomes tyrosine phosphorylated through the activation and association with the receptor of JAK2 and JAK3 kinases. After SDF-1α binding, JAK2 and JAK3 associate with CXCR4 and are activated, probably by transphosphorylation, in a Gαi-independent ...
TL;DR: Data suggest that in obesity‐related insulin resistance, the metabolic capacity of skeletal muscle appears to be organized toward fat esterification rather than oxidation and that dietary‐induced weight loss does not correct this disposition.
Abstract: A number of biochemical defects have been identified in glucose metabolism within skeletal muscle in obesity, and positive effects of weight loss on insulin resistance are also well established. Less is known about the capacity of skeletal muscle for the metabolism of fatty acids in obesity-related insulin resistance and of the effects of weight loss, though it is evident that muscle contains increased triglyceride. The current study was therefore undertaken to profile markers of human skeletal muscle for fatty acid metabolism in relation to obesity, in relation to the phenotype of insulin-resistant glucose metabolism, and to examine the effects of weight loss. Fifty-five men and women, lean and obese, with normal glucose tolerance underwent percutaneous biopsy of vastus lateralis skeletal muscle for determination of HADH, CPT, heparin-releasable (Hr) and tissue-extractable (Ext) LPL, CS, COX, PFK, and GAPDH enzyme activities, and content of cytosolic and plasma membrane FABP. Insulin sensitivity was meas...
TL;DR: There is a beneficial threshold of HO‐1 overexpression related to the accumulation of reactive iron released in the degradation of heme, which makes it unlikely that exaggerated expression ofHO‐1 is a cytoprotective response.
Abstract: It is often postulated that the cytoprotective nature of heme oxygenase (HO-1) explains the inducible nature of this enzyme. However, the mechanisms by which protection occurs are not verified by systematic evaluation of the physiological effects of HO. To explain how induction of HO-1 results in protection against oxygen toxicity, hamster fibroblasts (HA-1) were stably transfected with a tetracycline response plasmid containing the full-length rat HO-1 cDNA construct to allow for regulation of gene expression by varying concentrations of doxycycline (Dox). Transfected cells were exposed to hyperoxia (95% O2/5% CO2) for 24 h and several markers of oxidative injury were measured. With varying concentrations of Dox, HO activity was regulated between 3- and 17-fold. Despite cytoprotection with low (less than fivefold) HO activity, high levels of HO-1 expression (greater than 15-fold) were associated with significant oxygen cytotoxicity. Levels of non-heme reactive iron correlated with cellular injury in hype...
TL;DR: The DNA fragment encoding hepatitis B virus surface antigen was introduced into Agrobacterium tumerifacience LBA4404 and used to obtain transgenic lupin (Lupinus luteus L.) and lettuce (Lactuca sativa L.) cv and Burpee Bibb expressing envelope surface protein, which developed specific serum‐IgG response to plant produced protein.
Abstract: The infectious hepatitis B virus represents 42 nm spherical double-shelled particles. However, analysis of blood from hepatitis B virus carriers revealed the presence of smaller 22 nm particles consisting of a viral envelope surface protein. These particles are highly immunogenic and have been used in the design of hepatitis B virus vaccine produced in yeast. Upon expression in yeast, these proteins form virus-like particles that are used for parenteral immunization. Therefore, the DNA fragment encoding hepatitis B virus surface antigen was introduced into Agrobacterium tumerifacience LBA4404 and used to obtain transgenic lupin (Lupinus luteus L.) and lettuce (Lactuca sativa L.) cv. Burpee Bibb expressing envelope surface protein. Mice that were fed the transgenic lupin tissue developed significant levels of hepatitis B virus-specific antibodies. Human volunteers, fed with transgenic lettuce plants expressing hepatitis B virus surface antigen, developed specific serum-IgG response to plant produced protein.
TL;DR: Since haplogroups are defined on the basis of inherited variants, these data show that mtDNA inherited variability could play a role in successful aging and longevity.
Abstract: Mitochondrial DNA (mtDNA) is characterized by high variability, maternal inheritance, and absence of recombination. Studies of human populations have revealed ancestral associated polymorphisms whose combination defines groups of mtDNA types (haplogroups) that are currently used to reconstruct human evolution lineages. We used such inherited mtDNA markers to compare mtDNA population pools between a sample of individuals selected for successful aging and longevity (212 subjects older than 100 years and in good clinical condition) and a sample of 275 younger individuals (median age 38 years) carefully matched as to sex and geographic origin (northern and southern Italy). All nine haplogroups that are typical of Europeans were found in both samples, but male centenarians emerged in northern Italy as a particular sample: 1) mtDNA haplogroup frequency distribution was different between centenarians and younger individuals (P=0.017 by permutation tests); and 2) the frequency of the J haplogroup was notably high...
TL;DR: The insulin receptor is a transmembrane protein of the plasma membrane, where it recognizes extracellular insulin and transmits signals into the cellular signaling network, and is dependent on the caveolae environment for signaling.
Abstract: The insulin receptor is a transmembrane protein of the plasma membrane, where it recognizes extracellular insulin and transmits signals into the cellular signaling network. We report that insulin r ...
TL;DR: It is shown that reactions of the thiolactone with protein lysine residues were robust under physiological conditions, and protein homocysteinylation led to protein damage, manifested as multimerization and precipitation of extensively modified proteins.
Abstract: Homocysteine thiolactone, a cyclic thioester, is synthesized by certain aminoacyl-tRNA synthetases in editing or proofreading reactions that prevent translational incorporation of homocysteine into proteins. Although homocysteine thiolactone is expected to acylate amino groups in proteins, virtually nothing is known regarding reactivity of the thiolactone. Here it is shown that reactions of the thiolactone with protein lysine residues were robust under physiological conditions. In human serum incubated with homocysteine thiolactone, protein homocysteinylation was a major reaction that could be observed with as little as 10 nM thiolactone. Individual proteins were homocysteinylated at rates proportional to their lysine contents. Homocysteinylation led to protein damage, manifested as multimerization and precipitation of extensively modified proteins. Model enzymes, such as methionyl-tRNA synthetase and trypsin, were inactivated by homocysteinylation. Metabolic conversion of homocysteine to the thiolactone,...
TL;DR: It is shown that proliferating preadipocytes in cell lines and primary cultures, develop phagocytic activity toward microorganisms, and this function disappears when pre adipocytes stop proliferating and differentiate into adipocytes, raising the possibility of a potential direct involvement of adipose tissue in inflammatory processes.
Abstract: Several lines of evidence have supported a link betweeen adipose tissue and immunocompetent cells. This link is illustrated in obesity, where excess adiposity and impaired immune function have been described in both humans and genetically obese rodents. In addition, numerous factors involved in inflammatory response are secreted by both preadipocytes and macrophages. Here we show that proliferating preadipocytes in cell lines and primary cultures, develop phagocytic activity toward microorganisms. This is demonstrated by phagocytosis assays and confocal microscopy. This function disappears when preadipocytes stop proliferating and differentiate into adipocytes. After phagocytosis, preadipocytes show microbicide activity via an oxygen-dependent mechanism. In addition, preadipocytes as well as adipocytes are stained with MOMA-2, a marker of monocyte-macrophage lineage, but are negative for specific mature macrophage markers (F4/80 and Mac-1). These results suggest that preadipocytes could function as macrop...
TL;DR: The results suggest that the formation of reactive oxygen species (ROS) is a key mediator of cold‐induced apoptosis, with ROS formation being completely iron‐ mediated in liver endothelial cells and partially iron‐mediated in hepatocytes.
Abstract: When cultured hepatocytes were incubated in cell culture medium at 4°C for up to 30 h and then returned to 37°C, blebbing of the plasma membrane, cell detachment, chromatin condensation and margination, enhanced nuclear stainability with Hoechst 33342, ruffling of the nuclear membrane, and DNA fragmentation occurred. Similar to hepatocytes, cultured liver endothelial cells exhibited blebbing, chromatin condensation and margination, marked nuclear condensation, and increased stainability with Hoechst 33342 when exposed to hypothermia/rewarming. In both cell types, the occurrence and extent of these alterations were dependent on the duration of the cold incubation period. This cold-induced apoptosis was inhibited by hypoxia, by an array of free radical scavengers/antioxidants, and by iron chelators. However, the extent of the protection by the different antioxidants was different in the two cell types: iron chelators provided complete protection in liver endothelial cells but only partial protection in hepa...
TL;DR: The benefits of vitamin E supplementation alone or together with other antioxidants that work in concert with α‐TOH in ameliorating lipoprotein lipid peroxidation in the artery wall and, hence, atherosclerosis are discussed.
Abstract: The 'oxidation theory' of atherosclerosis proposes that oxidation of low density lipoprotein (LDL) contributes to atherogenesis. Although little direct evidence for a causative role of 'oxidized LDL' in atherogenesis exists, several studies show that, in vitro, oxidized LDL exhibits potentially proatherogenic activities and lipoproteins isolated from atherosclerotic lesions are oxidized. As a consequence, the molecular mechanisms of LDL oxidation and the actions of alpha-tocopherol (alpha-TOH, vitamin E), the major lipid-soluble lipoprotein antioxidant, have been studied in detail. Based on the known antioxidant action of alpha-TOH and epidemiological evidence, vitamin E is generally considered to be beneficial in coronary artery disease. However, intervention studies overall show a null effect of vitamin E on atherosclerosis. This confounding outcome can be rationalized by the recently discovered diverse role for alpha-TOH in lipoprotein oxidation; that is, alpha-TOH displays neutral, anti-, or, indeed, pro-oxidant activity under various conditions. This review describes the latter, novel action of alpha-TOH, termed tocopherol-mediated peroxidation, and discusses the benefits of vitamin E supplementation alone or together with other antioxidants that work in concert with alpha-TOH in ameliorating lipoprotein lipid peroxidation in the artery wall and, hence, atherosclerosis.
TL;DR: It is proposed that, considering the poor glucose control found in diabetics as well as the key role of oxidative stress in vascular complications, glycation‐mediated and free radical‐induced impairment of the antioxidant properties of albumin might be important parameters in vascular complication encountered in diabetes.
Abstract: Epidemiological data consistently show that reduced levels of serum albumin, which is the most abundant protein in plasma, are associated with an increased mortality risk. Various biological properties evidenced by direct effects of the albumin molecule may explain its beneficial effects. The present work aimed to investigate in vitro whether glycation or free radicals or both factors would affect the antioxidant properties of bovine serum albumin (BSA). Glycation was performed by long-term incubations (60 days) of BSA with increasing concentrations of glucose (up to 500 mmol/l) at 37°C. Minimally oxidized BSA was obtained after controlled incubations of dialyzed BSA samples with a water-soluble free radical generator [2,2′ azo-bis(2-amidinopropane) HCl]. The glycation-mediated modifications and the free radical-induced conformational changes of BSA were monitored using intrinsic fluorescence measurements of the tryptophan residues and acrylamide as a quenching agent. Thiol groups, Amadori glycophore cont...
TL;DR: (R)‐α‐Lipoic acid‐supplemented old rats have improved mitochondrial function, decreased oxidative damage, and increased metabolic rate, which improves indices of metabolic activity as well as lowers oxidative stress and damage evident in aging.
Abstract: A diet supplemented with (R)-lipoic acid, a mitochondrial coenzyme, was fed to old rats to determine its efficacy in reversing the decline in metabolism seen with age Young (3 to 5 months) and old (24 to 26 months) rats were fed an AIN-93M diet with or without (R)-lipoic acid (05% w/w) for 2 wk, killed, and their liver parenchymal cells were isolated Hepatocytes from untreated old rats vs young controls had significantly lower oxygen consumption (P<003) and mitochondrial membrane potential (R)-Lipoic acid supplementation reversed the age-related decline in O2 consumption and increased (P<003) mitochondrial membrane potential Ambulatory activity, a measure of general metabolic activity, was almost threefold lower in untreated old rats vs controls, but this decline was reversed (P<0005) in old rats fed (R)-lipoic acid The increase of oxidants with age, as measured by the fluorescence produced on oxidizing 2′,7′-dichlorofluorescin, was significantly lowered in (R)-lipoic acid supplemented old rats
TL;DR: It is reported here that mice lacking heart‐type FABP (H‐FABP) exhibit a severe defect of peripheral (non‐hepatic, non‐fat) LCFA utilization, establishing a requirement for H‐FabP in cardiac intracellular lipid transport and fuel selection and a major role in metabolic homeostasis.
Abstract: Nonenzymatic cytosolic fatty acid binding proteins (FABPs) are abundantly expressed in many animal tissues with high rates of fatty acid metabolism. No physiological role has been demonstrated for any FABP, although these proteins have been implicated in transport of free long-chain fatty acids (LCFAs) and protection against LCFA toxicity. We report here that mice lacking heart-type FABP (H-FABP) exhibit a severe defect of peripheral (nonhepatic, non-fat) LCFA utilization. In these mice, the heart is unable to efficiently take up plasma LCFAs, which are normally its main fuel, and switches to glucose usage. Altered plasma levels of LCFAs, glucose, lactate and beta-hydroxybutyrate are consistent with depressed peripheral LCFA utilization, intensified carbohydrate usage, and increased hepatic LCFA oxidation; these changes are most pronounced under conditions favoring LCFA oxidation. H-FABP deficiency is only incompletely compensated, however, causing acute exercise intolerance and, at old age, a localized cardiac hypertrophy. These data establish a requirement for H-FABP in cardiac intracellular lipid transport and fuel selection and a major role in metabolic homeostasis. This new animal model should be particularly useful for investigating the significance of peripheral LCFA utilization for heart function, insulin sensitivity, and blood pressure.
TL;DR: Analysis of the purified recombinant c‐Jun DNA binding domain for redox‐dependent thiol modifications and concomitant changes in DNA binding activity shows that changes in the ratio of reduced to oxidized glutathione provide the potential to oxidize c‐ Jun sulfhydryls by mechanisms that include both protein disulfide formation and S‐glutathiolation.
Abstract: Redox control of the transcription factor c-Jun maps to a single cysteine in its DNA binding domain. However, the nature of the oxidized state of this cysteine and, thus, the potential molecular mechanisms accounting for the redox regulation of c-Jun DNA binding remain unclear. To address this issue, we have analyzed the purified recombinant c-Jun DNA binding domain for redox-dependent thiol modifications and concomitant changes in DNA binding activity. We show that changes in the ratio of reduced to oxidized glutathione provide the potential to oxidize c-Jun sulfhydryls by mechanisms that include both protein disulfide formation and S-glutathiolation. We provide evidence that S-glutathiolation, which is specifically targeted to the cysteine residue located in the DNA binding site of the protein, may account for the reversible redox regulation of c-Jun DNA binding. Furthermore, based on a molecular model of the S-glutathiolated protein, we discuss the structural elements facilitating S-glutathiolation and...