BBA - Molecular Basis of Disease (v.1852, #9)
Editorial Board (i).
High-fat diet-induced deregulation of hippocampal insulin signaling and mitochondrial homeostasis deficiences contribute to Alzheimer disease pathology in rodents by Dmitry Petrov; Ignacio Pedrós; Gonzalo Artiach; Francesc X. Sureda; Emma Barroso; Mercè Pallàs; Gemma Casadesús; Carlos Beas-Zarate; Eva Carro; Isidro Ferrer; Manuel Vazquez-Carrera; Jaume Folch; Antoni Camins (1687-1699).
Global obesity is a pandemic status, estimated to affect over 2 billion people, that has resulted in an enormous strain on healthcare systems worldwide. The situation is compounded by the fact that apart from the direct costs associated with overweight pathology, obesity presents itself with a number of comorbidities, including an increased risk for the development of neurodegenerative disorders. Alzheimer disease (AD), the main cause of senile dementia, is no exception. Spectacular failure of the pharmaceutical industry to come up with effective AD treatment strategies is forcing the broader scientific community to rethink the underlying molecular mechanisms leading to cognitive decline. To this end, the emphasis is once again placed on the experimental animal models of the disease. In the current study, we have focused on the effects of a high-fat diet (HFD) on hippocampal-dependent memory in C57/Bl6 Wild-type (WT) and APPswe/PS1dE9 (APP/PS1) mice, a well-established mouse model of familial AD. Our results indicate that the continuous HFD administration starting at the time of weaning is sufficient to produce β-amyloid-independent, hippocampal-dependent memory deficits measured by a 2-object novel-object recognition test (NOR) in mice as early as 6 months of age. Furthermore, the resulting metabolic syndrome appears to have direct effects on brain insulin regulation and mitochondrial function. We have observed pathological changes related to both the proximal and distal insulin signaling pathway in the brains of HFD-fed WT and APP/PS1 mice. These changes are accompanied by a significantly reduced OXPHOS metabolism, suggesting that mitochondria play an important role in hippocampus-dependent memory formation and retention in both the HFD-treated and AD-like rodents at a relatively young age.Display Omitted
Keywords: APPSwe/PS1dE9; Insulin receptor; Mitochondria; Hippocampus; TAU; Alzheimer disease;
Mechanotransduction pathways in bone pathobiology by Anastasia Spyropoulou; Konstantinos Karamesinis; Efthimia K. Basdra (1700-1708).
The skeleton is subject to dynamic changes throughout life and bone remodeling is essential for maintenance of bone functionality. The cell populations which predominantly participate in bone and cartilage remodeling, namely osteocytes, osteoblasts, osteoclasts and chondrocytes sense and respond to external mechanical signals and via a series of molecular cascades control bone metabolism and turnover rate. The aforementioned process, known as mechanotransduction, is the underlying mechanism that controls bone homeostasis and function. A wide array of cross-talking signaling pathways has been found to play an important role in the preservation of bone and cartilage tissue health. Moreover, alterations in bone mechanotransduction pathways, due to genetic, hormonal and biomechanical factors, are considered responsible for the pathogenesis of bone and cartilage diseases. Extensive research has been conducted and demonstrated that aberrations in mechanotransduction pathways result in disease-like effects, however only few signaling pathways have actually been engaged in the development of bone disease. The aim of the present review is to present these signaling molecules and cascades that have been found to be mechano-responsive and implicated in bone disease development, as revealed by research in the last five years. In addition, the role of these molecules as prognostic or diagnostic disease markers and their potential as therapeutic targets are also discussed.
Keywords: Bone disease; Mechanotransduction; Osteoarthritis; Osteoporosis; Osteosarcoma; Chondrosarcoma;
Salusin-β contributes to vascular remodeling associated with hypertension via promoting vascular smooth muscle cell proliferation and vascular fibrosis by Hai-Jian Sun; Tong-Yan Liu; Feng Zhang; Xiao-Qing Xiong; Jue-Jin Wang; Qi Chen; Yue-Hua Li; Yu-Ming Kang; Ye-Bo Zhou; Ying Han; Xing-Ya Gao; Guo-Qing Zhu (1709-1718).
Vascular smooth muscle cell (VSMC) proliferation and vascular fibrosis are closely linked with hypertension and atherosclerosis. Salusin-β is a bioactive peptide involved in the pathogenesis of atherosclerosis. However, it is still largely undefined whether salusin-β is a potential candidate in the VSMC proliferation and vascular fibrosis. Experiments were carried out in human vascular smooth muscle cells (VSMCs) and in rats with intravenous injection of lentivirus expressing salusin-β. In vitro, salusin-β promoted VSMCs proliferation, which was attenuated by adenylate cyclase inhibitor SQ22536, PKA inhibitor Rp-cAMP, epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor AG1478, ERK inhibitor U0126 or cAMP response element binding protein (CREB) inhibitor KG501. It promoted the phosphorylation of ERK1/2, CREB and EGFR, which were abolished by SQ22536 or Rp-cAMP. Furthermore, epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor AG1478 diminished the salusin-β-evoked ERK1/2 and CREB phosphorylation. On the other hand, salusin-β increased collagen-I, collagen-III, fibronectin and connective tissue growth factor (CTGF) mRNA and phosphorylation of Smad2/3, which were prevented by ALK5 inhibitor A83-01. In vivo, salusin-β overexpression increased the media thickness, media/lumen ratio coupled with ERK1/2, CREB, EGFR and Smad2/3 phosphorylation, as well as the mRNA of collagen-I, collagen-III, fibronectin, transforming growth factor-β1 (TGF-β1) and CTGF in arteries. Moreover, salusin-β overexpression in rats caused severe hypertension. Intravenous injection of salusin-β dose-relatedly increased blood pressure, but excessive salusin-β decreased blood pressure and heart rate. These results indicate that salusin-β promotes VSMC proliferation via cAMP-PKA-EGFR-CREB/ERK pathway and vascular fibrosis via TGF-β1-Smad pathway. Increased salusin-β contributes to vascular remodeling and hypertension.Display Omitted
Keywords: Salusin; Vascular remodeling; Hypertension; Proliferation; Fibrosis;
Adenomatous polyposis coli (APC)-induced apoptosis of HT29 colorectal cancer cells depends on mitochondrial oxidative metabolism by Maricarmen Cristofaro; Annalisa Contursi; Simona D'Amore; Nicola Martelli; Ada Fiorenza Spaziante; Antonio Moschetta; Gaetano Villani (1719-1728).
Adenomatous polyposis coli (APC) is a tumor suppressor involved in the Wnt signaling, the primary driving force of the intestinal epithelium homeostasis. Alterations of components of the Wnt pathway, and in most cases mutations of APC, have been reported to promote colorectal cancer (CRC). During differentiation the enterocytes migrate from the crypt to the tip of the villus where they undergo apoptosis thus ensuring the continual renewal of the intestinal mucosa. The differentiation process is characterized by an activation gradient of the Wnt signaling pathway accompanied by a metabolic switch from glycolysis to mitochondrial oxidative phosphorylation along the crypt–villus axis. In the present work, we study the relationship between the expression of wild type APC protein and mitochondrial oxidative metabolism in HT29 colorectal cancer cells, originally carrying endogenous inactive APC alleles. By generating mtDNA-depleted (rho0) APC-inducible HT29 cells, we demonstrate for the first time that the APC-dependent apoptosis requires the production of reactive oxygen species (ROS) by the mitochondrial respiratory chain. The possible role of mitochondria as putative target in the prevention and/or therapy of colorectal cancer is herein discussed.
Keywords: Mitochondria; Respiratory chain; Reactive oxygen species (ROS); Wnt signaling; Apoptosis; Colorectal cancer;
Ink4/Arf locus restores glucose tolerance and insulin sensitivity by reducing hepatic steatosis and inflammation in mice with impaired IRS2-dependent signalling by Ángela Vinué; Irene Andrés-Blasco; Andrea Herrero-Cervera; Laura Piqueras; Vicente Andrés; Deborah J. Burks; María Jesús Sanz; Herminia González-Navarro (1729-1742).
Single nucleotide polymorphisms near the Ink4/Arf locus have been associated with type-2 diabetes mellitus. Previous studies indicate a protective role of the locus in the carbohydrate metabolism derangement associated with ageing in wild-type mice. The present study demonstrates that the increased Ink4/Arf locus expression in 1-year-old mice, partially-deficient for the insulin receptor substrate (IRS)2 (Irs2 +/− SuperInk4/Arf mice) ameliorates hepatic steatosis, inflammation and insulin resistance. Irs2 +/− SuperInk4/Arf mice displayed improved glucose tolerance and insulin sensitivity compared with Irs2 +/− mice which were glucose intolerant and insulin resistant compared with age-matched wild-type mice. These changes in Irs2 +/− mice were accompanied by enhanced hepatic steatosis, proinflammatory macrophage phenotype, increased Ly6Chi-monocyte percentage, T-lymphocyte activation and MCP1 and TNF-α cytokine levels. In Irs2 +/− SuperInk4/Arf mice, steatosis and inflammatory parameters were markedly reduced and similar to those of wild-type counterparts. In vivo insulin signalling also revealed reduced activation of the IRS/AKT-dependent signalling in Irs2 +/− mice. This was restored upon increased locus expression in Irs2 +/− SuperInk4/Arf mice which display similar activation levels as those for wild-type mice. In vivo treatment of Irs2 +/− SuperInk4/Arf mice with TNF-α diminished insulin canonical IRS/AKT-signalling and enhanced the stress SAPK/JNK–phosphoSer307IRS1-pathway suggesting that cytokine levels might potentially affect glucose homeostasis through changes in these insulin-signalling pathways. Altogether, these results indicate that enhanced Ink4/Arf locus expression restores glucose homeostasis and that this is associated with diminished hepatic steatosis and inflammation in mice with insulin resistance. Therefore, pharmacological interventions targeted to modulate the Ink4/Arf locus expression could be a tentative therapeutic approach to alleviate the inflammation associated with insulin resistance.
Keywords: CDKN2A/2B; Diabetes; Insulin resistance; Inflammation; Steatosis; Macrophage;
miRNA-584-5p exerts tumor suppressive functions in human neuroblastoma through repressing transcription of matrix metalloproteinase 14 by Xuan Xiang; Hong Mei; Hongxia Qu; Xiang Zhao; Dan Li; Huajie Song; Wanju Jiao; Jiarui Pu; Kai Huang; Liduan Zheng; Qiangsong Tong (1743-1754).
Matrix metalloproteinase 14 (MMP-14) is a membrane-anchored MMP crucial for tumorigenesis and aggressiveness, and is highly expressed in neuroblastoma (NB), the most common extracranial solid tumor in childhood. Recent evidence shows the emerging roles of endogenous promoter-targeting microRNAs (miRNAs) in regulating gene transcription. However, the roles of miRNAs in the transcription of MMP-14 still remain largely unknown. In this study, through mining computational algorithm program and Argonaute-chromosome interaction dataset, we identified one binding site of miRNA-584-5p (miR-584-5p) within the MMP-14 promoter. In NB tissues, miR-584-5p was under-expressed and inversely correlated with MMP-14 expression, and was an independent prognostic factor for favorable outcome of patients. miR-584-5p precursor attenuated the expression of MMP-14 in a Dicer-dependent manner, resulting in decreased levels of vascular endothelial growth factor, in cultured NB cell lines. In addition, miR-584-5p suppressed the promoter activity of MMP-14, and mutation of miR-584-5p binding site abolished these effects. Mechanistically, miR-584-5p recruited Argonaute 2 to facilitate the enrichment of enhancer of zeste homolog 2, histone H3 lysine 27 trimethylation, and histone H3 lysine 9 dimethylation on MMP-14 promoter in NB cells, which was abolished by repressing the miR-584-5p-promoter interaction. Gain- and loss-of-function studies demonstrated that miR-584-5p suppressed the growth, invasion, metastasis, and angiogenesis of NB cells in vitro and in vivo. Moreover, restoration of MMP-14 expression rescued the NB cells from changes in these biological features. Taken together, these results indicate that promoter-targeting miR-584-5p exerts tumor suppressive functions in NB through repressing the transcription of MMP-14.
Keywords: Neuroblastoma; microRNA-584-5p; Matrix metalloproteinase 14; Transcriptional repression;
Neuraminidase-1 mediates skeletal muscle regeneration by Juliana de Carvalho Neves; Vanessa Rodrigues Rizzato; Alan Fappi; Mariana Miranda Garcia; Gerson Chadi; Diantha van de Vlekkert; Alessandra d'Azzo; Edmar Zanoteli (1755-1764).
Neuraminidase-1 (NEU1) is the sialidase responsible for the catabolism of sialoglycoconjugates in lysosomes. Congenital NEU1 deficiency causes sialidosis, a severe lysosomal storage disease associated with a broad spectrum of clinical manifestations, which also include skeletal deformities, skeletal muscle hypotonia and weakness. Neu1−/− mice, a model of sialidosis, develop an atypical form of muscle degeneration caused by progressive expansion of the connective tissue that infiltrates the muscle bed, leading to fiber degeneration and atrophy. Here we investigated the role of Neu1 in the myogenic process that ensues during muscle regeneration after cardiotoxin-induced injury of limb muscles. A comparative analysis of cardiotoxin-treated muscles from Neu1−/− mice and Neu1+/+ mice showed increased inflammatory and proliferative responses in the absence of Neu1 during the early stages of muscle regeneration. This was accompanied by significant and sequential upregulation of Pax7, MyoD, and myogenin mRNAs. The levels of both MyoD and myogenin proteins decreased during the late stages of regeneration, which most likely reflected an increased rate of degradation of the myogenic factors in the Neu1−/− muscle. We also observed a delay in muscle cell differentiation, which was characterized by prolonged expression of embryonic myosin heavy chain, as well as reduced myofiber cross-sectional area. At the end of the regenerative process, collagen type III deposition was increased compared to wild-type muscles and internal controls, indicating the initiation of fibrosis. Overall, these results point to a role of Neu1 throughout muscle regeneration.
Keywords: NEU1; Sialidosis; Skeletal muscle regeneration; Cell proliferation; Muscle maturation; Fibrosis;
Recent insights on the role of cholesterol in non-alcoholic fatty liver disease by Graciela Arguello; Elisa Balboa; Marco Arrese; Silvana Zanlungo (1765-1778).
Non-alcoholic fatty liver disease (NAFLD) encompasses a spectrum of hepatic histopathological changes ranging from non-inflammatory intracellular fat deposition to non-alcoholic steatohepatitis (NASH), which may progress into hepatic fibrosis, cirrhosis, or hepatocellular carcinoma. NAFLD hallmark is the excessive hepatic accumulation of neutral lipids that result from an imbalance between lipid availability and lipid removal. Recent data suggest that disturbed hepatic cholesterol homeostasis and liver free cholesterol (FC) accumulation are relevant to the pathogenesis of NAFLD/NASH. Hepatic FC accumulation in NAFLD results from alterations in intracellular cholesterol transport and from unbalanced cellular cholesterol homeostasis characterized by activation of cholesterol biosynthetic pathways, increased cholesterol de-esterification and attenuation of cholesterol export and bile acid synthesis pathways. FC accumulation leads to liver injury through the activation of intracellular signaling pathways in Kupffer cells (KCs), Stellate cells (HSCs) and hepatocytes. The activation of KCs and HSCs promotes inflammation and fibrogenesis. In addition, FC accumulation in liver mitochondria induces mitochondrial dysfunction, which results in increasing production of reactive oxygen species, and triggers the unfolded protein response in the endoplasmic reticulum (ER) causing ER stress and apoptosis. These events create a vicious circle that contributes to the maintenance of steatosis and promotes ongoing hepatocyte death and liver damage, which in turn may translate into disease progression. In the present review we summarize the current knowledge on dysregulated cholesterol homeostasis in NAFLD and examine the cellular mechanisms of hepatic FC toxicity and its contribution to ongoing liver injury in this disease. The therapeutic implications of this knowledge are also discussed.Display Omitted
Keywords: NAFLD; NASH; Lipotoxicity; Free cholesterol; Cholesterol homeostasis; Liver;
Gliadin-mediated production of polyamines by RAW264.7 macrophages modulates intestinal epithelial permeability in vitro by Amelia Barilli; Bianca Maria Rotoli; Rossana Visigalli; Filippo Ingoglia; Martina Cirlini; Barbara Prandi; Valeria Dall'Asta (1779-1786).
Celiac disease (CD) is an immune-mediated enteropathy sustained by dietary gluten in susceptible individuals, and characterized by a complex interplay between adaptive and innate responses against gluten peptides (PTG). In a recent contribution we have demonstrated that the treatment with PTG induces the expression and activity of arginase in both murine macrophages and human monocytes from healthy subjects, thus suggesting a role for arginine and its metabolites in gluten-triggered response of these cells. Here we further explore this field, by addressing the effects of PTG on polyamine synthesis and release in murine RAW264.7 macrophages, and how they affect epithelial permeability of Caco-2 monolayers. Results obtained show a massive production and release of putrescine by macrophages upon incubation with gluten peptides; this, in turn, causes a decrease in TEER in epithelial cells, indicating that PTG-driven secretion of polyamines by macrophages has a role in the modulation of intestinal permeability in vitro. At a molecular level, putrescine production appears referable to the activation of C/EBPβ transcription factor, which is known to be responsible for arginase induction in activated macrophages and is a crucial mediator of inflammation. Whether these pathways are stimulated also in vivo deserves to be further investigated, as well as their role in gluten-driven cellular and intestinal defects typical of CD patients.
Keywords: Celiac disease; Gliadin; Polyamines; Arginase; Intestinal permeability;
Mechanism for increased hepatic glycerol synthesis in the citrin/mitochondrial glycerol-3-phosphate dehydrogenase double-knockout mouse: Urine glycerol and glycerol 3-phosphate as potential diagnostic markers of human citrin deficiency by Mitsuaki Moriyama; Yuki Fujimoto; Shizuka Rikimaru; Miharu Ushikai; Eishi Kuroda; Kenji Kawabe; Katsura Takano; Akihiro Asakawa; Akio Inui; Kazuhiro Eto; Takashi Kadowaki; David S. Sinasac; Yoshiyuki Okano; Masahide Yazaki; Shu-ichi Ikeda; Chunhua Zhang; Yuan-Zong Song; Osamu Sakamoto; Shigeo Kure; Hiroshi Mitsubuchi; Fumio Endo; Masahisa Horiuchi; Yoichi Nakamura; Ken-ichi Yamamura; Takeyori Saheki (1787-1795).
The mitochondrial aspartate-glutamate carrier isoform 2 (citrin) and mitochondrial glycerol-3-phosphate dehydrogenase (mGPD) double-knockout mouse has been a useful model of human citrin deficiency. One of the most prominent findings has been markedly increased hepatic glycerol 3-phosphate (G3P) following oral administration of a sucrose solution. We aimed to investigate whether this change is detectable outside of the liver, and to explore the mechanism underlying the increased hepatic G3P in these mice. We measured G3P and its metabolite glycerol in plasma and urine of the mice under various conditions. Glycerol synthesis from fructose was also studied using the liver perfusion system. The citrin/mGPD double-knockout mice showed increased urine G3P and glycerol under normal, fed conditions. We also found increased plasma glycerol under fasted conditions, while oral administration of different carbohydrates or ethanol led to substantially increased plasma glycerol. Fructose infusion to the perfused liver of the double-knockout mice augmented hepatic glycerol synthesis, and was accompanied by a concomitant increase in the lactate/pyruvate (L/P) ratio. Co-infusion of either pyruvate or phenazine methosulfate, a cytosolic oxidant, with fructose corrected the high L/P ratio, leading to reduced glycerol synthesis. Overall, these findings suggest that hepatic glycerol synthesis is cytosolic NADH/NAD+ ratio-dependent and reveal a likely regulatory mechanism for hepatic glycerol synthesis following a high carbohydrate load in citrin-deficient patients. Therefore, urine G3P and glycerol may represent potential diagnostic markers for human citrin deficiency.
Keywords: Citrin deficiency; Glycerol; Glycerol 3-phosphate; Mitochondrial aspartate-glutamate carrier; Redox state; Pyruvate;
Crosstalk between calpain activation and TGF-β1 augments collagen-I synthesis in pulmonary fibrosis by Feng-Zhi Li; Peng-Cheng Cai; Lin-Jie Song; Li-Ling Zhou; Qian Zhang; Shan-Shan Rao; Yu Xia; Fei Xiang; Jian-Bao Xin; Peter A. Greer; Huan-Zhong Shi; Yunchao Su; Wan-Li Ma; Hong Ye (1796-1804).
Idiopathic pulmonary fibrosis (IPF) is a chronic progressive lung disease of unknown cause that typically leads to respiratory failure and death within 3–5 years of diagnosis. TGF-β1 is considered a major profibrotic factor. However, TGF-β1 is necessary but not sufficient to the pathogenesis of fibrotic lesion of the lungs. Recent observations have revealed that calpain, a calcium dependent protease, plays a pivotal role in tissue remodeling and fibrosis. However, the mechanism of calpain mediating pulmonary fibrosis is not understood. Calpain conditional knockout (ER-Cre+/− capns1 flox/flox) mice and primary human lung fibroblasts (HLFs) were used here to investigate the relationship between calpain and TGF-β1. Calpain knockout mice were protected from fibrotic effects of bleomycin. Bleomycin induced increases in TGF-β1 via calpain activation in HLFs. Moreover, TGF-β1 also activated calpain. This crosstalk between calpain activation and TGF-β1 triggered the downstream signaling pathway including TGF-β1 Smad2/3 and non-Smad (Akt) pathways, as well as collagen-I synthesis. Taken together, our data indicate that the crosstalk between calpain activation and TGF-β1 augments collagen-I synthesis in HLFs and in pulmonary fibrosis. Intervention in the crosstalk between calpain activation and TGF-β1 is a novel potential strategy to prevent pulmonary fibrosis.
Keywords: Calpain; TGF-β1; Collagen; Pulmonary fibrosis; Fibroblasts; Smad;
Mitochondrial DNA: A disposable genome? by Inna N. Shokolenko; Mikhail F. Alexeyev (1805-1809).
In mammalian cells, mitochondria are the only organelles besides the nucleus that house genomic DNA. The mammalian mitochondrial genome is represented by prokaryotic-type, circular, highly compacted DNA molecules. Today, more than a half-century after their discovery, the biology of these small and redundant molecules remains much less understood than that of their nuclear counterparts. One peculiarity of the mitochondrial genome that emerged in recent years is its disposable nature, as evidenced by cells abandoning a fraction of their mitochondrial DNA (mtDNA) in response to various stimuli with little or no physiological consequence. Here, we review some recent developments in the field of mtDNA biology and discuss emerging questions on the disposability and indispensability of mtDNA.
Keywords: Mitochondrial DNA; mtDNA maintenance; mtDNA copy number; mtDNA degradation; Extramitohondrial mtDNA;
Synergistic effects of β-amyloid and ceramide-induced insulin resistance on mitochondrial metabolism in neuronal cells by Bumsup Kwon; Timothy Gamache; Han-Kyu Lee; Henry W. Querfurth (1810-1823).
A large body of evidence support major roles of mitochondrial dysfunction and insulin action in the Alzheimer's disease (AD) brain. However, interaction between cellular expression of β-amyloid (Aβ) and insulin resistance on mitochondrial metabolism has not been explored in neuronal cells. We investigated the additive and synergistic effects of intracellular Aβ42 and ceramide-induced insulin resistance on mitochondrial metabolism in SH-SY5Y and Neuro-2a cells. In our model, mitochondria take-up Aβ42 expressed through viral-mediated transfection and exposure of the same cells to ceramide produces resistance to insulin signaling. Ceramide alone increased phosphorylated MAP kinases while decreasing phospho-Akt (Ser473). The combination of Aβ42 and ceramide synergistically decreased phospho-Thr308 on Akt. Aβ42 and ceramide synergistically also decreased mitochondrial complex III activity and ATP generation whereas Aβ alone was largely responsible for complex IV inhibition and increases in mitochondrial reactive oxygen species production (ROS). Proteomic analysis showed that a number of mitochondrial respiratory chain and tricarboxylic acid cycle enzymes were additively or synergistically decreased by ceramide in combination with Aβ42 expression. Mitochondrial fusion and fission proteins were notably dysregulated by Aβ42 (Mfn1) or Aβ42 plus ceramide (OPA1, Drp1). Antioxidant vitamins blocked the Aβ42 alone-induced ROS production, but did not reverse Aβ42-induced ATP reduction or complex IV inhibition. Aβ expression combined with ceramide exposure had additive effects to decrease cell viability. Taken together, our data demonstrate that Aβ42 expression and ceramide-induced insulin resistance synergistically interact to exacerbate mitochondrial damage and that therapeutic efforts to reduce insulin resistance could lessen failures of energy production and mitochondrial dynamics.
Keywords: β-amyloid; Ceramide; Insulin resistance; Mitochondrial dysfunction; Alzheimer's disease;
Leptin modulates human Sertoli cells acetate production and glycolytic profile: a novel mechanism of obesity-induced male infertility? by Ana D. Martins; Ana C. Moreira; Rosália Sá; Mariana P. Monteiro; Mário Sousa; Rui A. Carvalho; Branca M. Silva; Pedro F. Oliveira; Marco G. Alves (1824-1832).
Human feeding behavior and lifestyle are gradually being altered, favoring the development of metabolic diseases, particularly type 2 diabetes and obesity. Leptin is produced by the adipose tissue acting as a satiety signal. Its levels have been positively correlated with fat mass and hyperleptinemia has been proposed to negatively affect male reproductive function. Nevertheless, the molecular mechanisms by which this hormone affects male fertility remain unknown. Herein, we hypothesize that leptin acts on human Sertoli cells (hSCs), the “nurse cells” of spermatogenesis, altering their metabolism. To test our hypothesis, hSCs were cultured without or with leptin (5, 25 and 50 ng/mL). Leptin receptor was identified by qPCR and Western blot. Protein levels of glucose transporters (GLUT1, GLUT2 and GLUT3), phosphofructokinase, lactate dehydrogenase (LDH) and monocarboxylate transporter 4 (MCT4) were determined by Western Blot. LDH activity was assessed and metabolite production/consumption determined by proton nuclear magnetic resonance. Oxidative damage was evaluated by assessing lipid peroxidation, protein carbonilation and nitration. Our data shows that leptin receptor is expressed in hSCs. The concentration of leptin found in lean, healthy patients, upregulated GLUT2 protein levels and concentrations of leptin found in lean and obese patients increased LDH activity. Of note, all leptin concentrations decreased hSCs acetate production illustrating a novel mechanism for this hormone action. Moreover, our data shows that leptin does not induce or protect hSCs from oxidative damage. We report that this hormone modulates the nutritional support of spermatogenesis, illustrating a novel mechanism that may be linked to obesity-induced male infertility.
Keywords: Sertoli cells; Leptin; Obesity; Spermatogenesis; Male fertility;
UV-induced retinal proteome changes in the rat model of age-related macular degeneration by Sandra Kraljević Pavelić; Marko Klobučar; Mirela Sedić; Vedran Micek; Peter Gehrig; Jonas Grossman; Krešimir Pavelić; Božidar Vojniković (1833-1845).
Age-related macular degeneration (AMD) is characterized by irreversible damage of photoreceptors in the central posterior part of the retina, called the macula and is the most common cause of vision loss in those aged over 50. A growing body of evidence shows that cumulative long-term exposure to UV radiation may be harmful to the retina and possibly leads to AMD irrespective of age. In spite of many research efforts, cellular and molecular mechanisms leading to UV-induced retinal damage and possibly retinal diseases such as AMD are not completely understood. In the present study we explored damage mechanisms accounting for UV-induced retinal phototoxicity in the rats exposed to UVA and UVB irradiation using a proteomics approach. Our study showed that UV irradiation induces profound changes in the retinal proteomes of the rats associated with the disruption of energy homeostasis, oxidative stress, DNA damage response and structural and functional impairments of the interphotoreceptor matrix components and their cell surface receptors such as galectins. Two small leucine-rich proteoglycans, biglycan and lumican, were identified as phototoxicity biomarkers associated with UV-induced disruption of interphotoreceptor matrix (IPM). In addition, UVB induced activation of Src kinase, which could account for cytoskeletal rearrangements in the retina was observed at the proteomics level. Pharmacological intervention either to target Src kinase with the aim of preventing cytoskeletal rearrangements in the retinal pigment epithelium (RPE) and neuronal retina or to help rebuild damaged IPM may provide fresh avenues of treatment for patients suffering from AMD.
Keywords: Age-related macular degeneration; UV irradiation; Oxidative stress; Interphotoreceptor matrix; Retinal pigment epithelium; Proteomics;
Kaiso overexpression promotes intestinal inflammation and potentiates intestinal tumorigenesis in ApcMin/+ mice by Christina C. Pierre; Joseph Longo; Meaghan Mavor; Snezana B. Milosavljevic; Roopali Chaudhary; Ebony Gilbreath; Clayton Yates; Juliet M. Daniel (1846-1855).
Constitutive Wnt/β-catenin signaling is a key contributor to colorectal cancer (CRC). Although inactivation of the tumor suppressor adenomatous polyposis coli (APC) is recognized as an early event in CRC development, it is the accumulation of multiple subsequent oncogenic insults facilitates malignant transformation. One potential contributor to colorectal carcinogenesis is the POZ-ZF transcription factor Kaiso, whose depletion extends lifespan and delays polyp onset in the widely used ApcMin/+ mouse model of intestinal cancer. These findings suggested that Kaiso potentiates intestinal tumorigenesis, but this was paradoxical as Kaiso was previously implicated as a negative regulator of Wnt/β-catenin signaling. To resolve Kaiso's role in intestinal tumorigenesis and canonical Wnt signaling, we generated a transgenic mouse model (KaisoTg/+ ) expressing an intestinal-specific myc-tagged Kaiso transgene. We then mated KaisoTg/+ and ApcMin/+ mice to generate KaisoTg/+ :ApcMin/+ mice for further characterization. KaisoTg/+ :ApcMin/+ mice exhibited reduced lifespan and increased polyp multiplicity compared to ApcMin/+ mice. Consistent with this murine phenotype, we found increased Kaiso expression in human CRC tissue, supporting a role for Kaiso in human CRC. Interestingly, Wnt target gene expression was increased in KaisoTg/+ :ApcMin/+ mice, suggesting that Kaiso's function as a negative regulator of canonical Wnt signaling, as seen in Xenopus, is not maintained in this context. Notably, KaisoTg/+ :ApcMin/+ mice exhibited increased inflammation and activation of NFκB signaling compared to their ApcMin/+ counterparts. This phenotype was consistent with our previous report that KaisoTg/+ mice exhibit chronic intestinal inflammation. Together our findings highlight a role for Kaiso in promoting Wnt signaling, inflammation and tumorigenesis in the mammalian intestine.
Keywords: Kaiso; Apc; Wnt; Tumorigenesis; Inflammation;
Epilepsy-causing mutations in Kv7.2 C-terminus affect binding and functional modulation by calmodulin by Paolo Ambrosino; Alessandro Alaimo; Silvia Bartollino; Laura Manocchio; Michela De Maria; Ilaria Mosca; Carolina Gomis-Perez; Araitz Alberdi; Giovanni Scambia; Gaetan Lesca; Alvaro Villarroel; Maurizio Taglialatela; Maria Virginia Soldovieri (1856-1866).
Mutations in the KCNQ2 gene, encoding for voltage-gated Kv7.2 K+ channel subunits, are responsible for early-onset epileptic diseases with widely-diverging phenotypic presentation, ranging from Benign Familial Neonatal Seizures (BFNS) to epileptic encephalopathy. In the present study, Kv7.2 BFNS-causing mutations (W344R, L351F, L351V, Y362C, and R553Q) have been investigated for their ability to interfere with calmodulin (CaM) binding and CaM-induced channel regulation. To this aim, semi-quantitative (Far-Western blotting) and quantitative (Surface Plasmon Resonance and dansylated CaM fluorescence) biochemical assays have been performed to investigate the interaction of CaM with wild-type or mutant Kv7.2 C-terminal fragments encompassing the CaM-binding domain; in parallel, mutation-induced changes in CaM-dependent Kv7.2 or Kv7.2/Kv7.3 current regulation were investigated by patch-clamp recordings in Chinese Hamster Ovary (CHO) cells co-expressing Kv7.2 or Kv7.2/Kv7.3 channels and CaM or CaM1234 (a CaM isoform unable to bind Ca2+). The results obtained suggest that each BFNS-causing mutation prompts specific biochemical and/or functional consequences; these range from slight alterations in CaM affinity which did not translate into functional changes (L351V), to a significant reduction in the affinity and functional modulation by CaM (L351F, Y362C or R553Q), to a complete functional loss without significant alteration in CaM affinity (W344R). CaM overexpression increased Kv7.2 and Kv7.2/Kv7.3 current levels, and partially (R553Q) or fully (L351F) restored normal channel function, providing a rationale pathogenetic mechanism for mutation-induced channel dysfunction in BFNS, and highlighting the potentiation of CaM-dependent Kv7.2 modulation as a potential therapeutic approach for Kv7.2-related epilepsies.
Keywords: Calmodulin; Kv7.2; Epilepsy; Surface Plasmon Resonance; Far-Western blotting; Fluorescence; Electrophysiology;
FNDC5 overexpression and irisin ameliorate glucose/lipid metabolic derangements and enhance lipolysis in obesity by Xiao-Qing Xiong; Dan Chen; Hai-Jian Sun; Lei Ding; Jue-Jin Wang; Qi Chen; Yue-Hua Li; Ye-Bo Zhou; Ying Han; Feng Zhang; Xing-Ya Gao; Yu-Ming Kang; Guo-Qing Zhu (1867-1875).
Irisin is a cleaved and secreted fragment of fibronectin type III domain containing 5 (FNDC5), and contributes to the beneficial effects of exercise on metabolism. Here we report the therapeutical effects of FNDC5/irisin on metabolic derangements and insulin resistance in obesity, and show the lipolysis effect of irisin and its signal molecular mechanism. In obese mice, lentivirus mediated-FNDC5 overexpression enhanced energy expenditure, lipolysis and insulin sensitivity, and reduced hyperlipidemia, hyperglycemia, hyperinsulinism, blood pressure and norepinephrine levels; it increased hormone-sensitive lipase (HSL) expression and phosphorylation, and reduced perilipin level and adipocyte diameter in adipose tissues. Subcutaneous perfusion of irisin reduced hyperlipidemia and hyperglycemia, and improved insulin resistance. Either FNDC5 overexpression or irisin perfusion only induced a tendency toward a slight decrease in body weight in obese mice. In 3T3-L1 adipocytes, irisin enhanced basal lipolysis rather than isoproterenol-induced lipolysis, which were prevented by inhibition of adenylate cyclase or PKA; irisin increased the HSL and perilipin phosphorylation; it increased PKA activity, and cAMP and HSL mRNA levels, but reduced perilipin expression. These results indicate that FNDC5/irisin ameliorates glucose/lipid metabolic derangements and insulin resistance in obese mice, and enhances lipolysis via cAMP–PKA–HSL/perilipin pathway. FNDC5 or irisin can be taken as an effective therapeutic strategy for metabolic disorders.Display Omitted
Keywords: Irisin; Obesity; Metabolism; Insulin resistance; Lipolysis;
NDRG1 attenuates epithelial–mesenchymal transition of nasopharyngeal cancer cells via blocking Smad2 signaling by Zhi-Yan Hu; Wei-Bing Xie; Fang Yang; Li-Wei Xiao; Xiao-Yan Wang; Shi-You Chen; Zu-Guo Li (1876-1886).
N-myc downstream-regulated gene 1 (NDRG1) has been implicated in tumorigenesis and metastasis in different cancers. However, its role in nasopharyngeal carcinoma remains unknown. We found that NDRG1 expression level was high in nasopharyngeal cancer 5-8F cells but low in 5-8F-LN cells with lymphatic metastasis potential. Knockdown of NDRG1 by shRNA promoted 5-8F cell proliferation, migration, and invasion in vitro and its tumorigenesis in vivo. Moreover, NDRG1 deficiency induced an epithelial–mesenchymal transition (EMT) of 5-8F cells as shown by an attenuation of E-cadherin and an induction of N-cadherin and vimentin expression. NDRG1 knockdown also enhanced Smad2 expression and phosphorylation. Smad2 signaling was attenuated in 5-8F cells but was significantly activated in 5-8F-LN cells. Knockdown of Smad2 restored E-cadherin but attenuated N-cadherin expression in NDRG1-deficient 5-8F cells, suggesting a reduction of EMT. Consistently, blockade of Smad2 in 5-8F-LN cells increased E-cadherin while diminishing N-cadherin and vimentin expression. These data indicate that Smad2 mediates the NDRG1 deficiency-induced EMT of 5-8F cells. In tumors derived from NDRG1-deficient 5-8F cells, E-cadherin expression was inhibited while vimentin and Smad2 were increased in a large number of cancer cells. Most importantly, NDRG1 expression was attenuated in human nasopharyngeal carcinoma tissues, resulted in a lower survival rate in patients. The NDRG1 was further decreased in the detached nasopharyngeal cancer cells, which was associated with a further reduced survival rate in patients with lymphatic metastasis. Taken together, these results demonstrated that NDRG1 prevents nasopharyngeal tumorigenesis and metastasis via inhibiting Smad2-mediated EMT of nasopharyngeal cells.
Keywords: N-myc downstream-regulated gene 1; Nasopharyngeal carcinoma; Proliferation; Metastasis; Epithelial–mesenchymal transition;
The emerging role of adiponectin in cerebrovascular and neurodegenerative diseases by Yang Yang; Wei Hu; Shuai Jiang; Bodong Wang; Yue Li; Chongxi Fan; Shouyin Di; Zhiqiang Ma; Wayne Bond Lau; Yan Qu (1887-1894).
Adiponectin is an anti-atherogenic protein secreted by adipose cells that improves insulin sensitivity. Notably, adiponectin receptors are expressed in the brain, suggesting that adiponectin signaling disruption may impact neurologic function. Recently, studies have demonstrated the association of adiponectin levels with cerebrovascular disorders and neurodegenerative diseases (NDDs), and these results have drawn significant attention. In this review, we discuss the association between the adiponectin levels and the incidence, progression, and prognosis of cerebrovascular disorders and NDDs. We describe the controversial issues surrounding current studies and present our hypothesis concerning the possible mechanism underlying adiponectin function in neurological disorders. Finally, we explicate obstacles preventing clinical adiponectin administration, including available routes of drug delivery and the central nervous system regulation of adiponectin. Collectively, the data assembled herein serve as a comprehensive reference regarding the role of adiponectin in neurological disorders to support the future clinical potential of adiponectin as a therapeutic agent.
Keywords: Adiponectin; Cerebrovascular diseases; Neurodegenerative diseases;
C/EBP homologous protein (CHOP) gene deficiency attenuates renal ischemia/reperfusion injury in mice by Mi Ra Noh; Jee In Kim; Sang Jun Han; Tae-Jin Lee; Kwon Moo Park (1895-1901).
C/EBP homologous protein (CHOP), a transcription factor for the expression of apoptosis-related genes, plays an important role in endoplasmic reticulum (ER) stress-related organ diseases, including diseases of the kidney. Here, we investigated the role of CHOP in ischemia/reperfusion (I/R)-induced acute kidney injury using CHOP-knockout (CHOP−/−) and wild type (CHOP+/+) mice. Fifteen or thirty minutes of bilateral renal ischemia (I/R) insult resulted in necrotic and apoptotic tubular epithelial cell death, together with increases in plasma creatinine (PCr) and blood urea nitrogen (BUN) concentrations. After I/R, BiP/GRP78 and CHOP expressions in the kidney gradually increased over time. CHOP expression was greater in the outer medulla than that in the cortex and localized intensely in the nucleus. I/R caused apoptosis of tubular epithelial cells in both CHOP−/− and CHOP+/+ mice. The number of apoptotic cells after I/R was lower in CHOP−/− mice than that in CHOP+/+ mice. Consistent with the degree of apoptosis, I/R-induced kidney morphological and functional damages were milder in CHOP−/− than that in CHOP+/+ mice. The cleavage of procaspase-3 and the induction of Bax protein after I/R were lower in CHOP−/− than that in CHOP+/+ mice. In contrast, the expression levels of Bcl-2, Bcl-xL, cIAP2, Mcl-1, and XIAP were higher in CHOP−/− than that in CHOP+/+ mice. These results indicate that I/R induces ER stress, leading to the activation of CHOP-associated apoptosis signals, resulting in renal functional and histological damages.
Keywords: C/EBP homologous protein; CHOP; ER stress; Ischemia; Apoptosis;
Ischemia-induced autophagy contributes to neurodegeneration in cerebellar Purkinje cells in the developing rat brain and in primary cortical neurons in vitro by Alicia K. Au; Yaming Chen; Lina Du; Craig M. Smith; Mioara D. Manole; Sirine A. Baltagi; Charleen T. Chu; Rajesh K. Aneja; Hülya Bayır; Patrick M. Kochanek; Robert S.B. Clark (1902-1911).
Increased autophagy/mitophagy is thought to contribute to cerebellar dysfunction in Purkinje cell degeneration mice. Intriguingly, cerebellar Purkinje cells are highly vulnerable to hypoxia–ischemia (HI), related at least in part to their high metabolic activity. Whether or not excessive or supraphysiologic autophagy plays a role in Purkinje cell susceptibility to HI is unknown. Accordingly, we evaluated the role of autophagy in the cerebellum after global ischemia produced by asphyxial cardiac arrest in postnatal day (PND) 16–18 rats, using siRNA-targeted inhibition of Atg7, necessary for microtubule-associated protein light chain 3-II (LC3-II) and Atg12–Atg5 complex formation. Two days before a 9 min asphyxial cardiac arrest or sham surgery, Atg7 or control siRNA was injected intracisternally to target the cerebellum. Treatment with Atg7 siRNA: 1) reduced Atg7 protein expression in the cerebellum by 56%; 2) prevented the typical ischemia-induced formation of LC3-II in the cerebellum 24 h after asphyxial cardiac arrest; 3) improved performance on the beam-balance apparatus on days 1–5; and 4) increased calbindin-labeled Purkinje cell survival assessed on day 14. Improved Purkinje cell survival was more consistent in female vs. male rats, and improved beam-balance performance was only seen in female rats. Similar responses to Atg7 siRNA i.e. reduced autophagy and neurodegeneration vs. control siRNA were seen when exposing sex-segregated green fluorescent protein-LC3 tagged mouse primary cortical neurons to oxygen glucose deprivation in vitro. Thus, inhibition of autophagy after global ischemia in PND 16–18 rats leads to increased survival of Purkinje cells and improved motor performance in a sex-dependent manner.
Keywords: Asphyxia; Atg7; Cardiac arrest; Cerebellum; Hypoxia–ischemia; Ischemic brain injury; Oxygen glucose deprivation; Purkinje neuron; Small interfering RNA;
Constitutive activities of estrogen-related receptors: Transcriptional regulation of metabolism by the ERR pathways in health and disease by Janice M. Huss; Wojciech G. Garbacz; Wen Xie (1912-1927).
The estrogen-related receptors (ERRs) comprise a small group of orphan nuclear receptor transcription factors. The ERRα and ERRγ isoforms play a central role in the regulation of metabolic genes and cellular energy metabolism. Although less is known about ERRβ, recent studies have revealed the importance of this isoform in the maintenance of embryonic stem cell pluripotency. Thus, ERRs are essential to many biological processes. The development of several ERR knockout and overexpression models and the application of advanced functional genomics have allowed rapid advancement of our understanding of the physiology regulated by ERR pathways. Moreover, it has enabled us to begin to delineate the distinct programs regulated by ERRα and ERRγ that have overlapping effects on metabolism and growth. The current review primarily focuses on the physiologic roles of ERR isoforms related to their metabolic regulation; therefore, the ERRα and ERRγ are discussed in the greatest detail. We emphasize findings from gain- and loss-of-function models developed to characterize ERR control of skeletal muscle, heart and musculoskeletal physiology. These models have revealed that coordinating metabolic capacity with energy demand is essential for seemingly disparate processes such as muscle differentiation and hypertrophy, innate immune function, thermogenesis, and bone remodeling. Furthermore, the models have revealed that ERRα- and ERRγ-deficiency in mice accelerates progression of pathologic processes and implicates ERRs as etiologic factors in disease. We highlight the human diseases in which ERRs and their downstream metabolic pathways are perturbed, including heart failure and diabetes. While no natural ligand has been identified for any of the ERR isoforms, the potential for using synthetic small molecules to modulate their activity has been demonstrated. Based on our current understanding of their transcriptional mechanisms and physiologic relevance, the ERRs have emerged as potential therapeutic targets for treatment of osteoporosis, muscle atrophy, insulin resistance and heart failure in humans.
Keywords: Nuclear hormone receptors; Orphan receptors; Energy metabolism; Mitochondria; Transcriptional regulation; Genetic mouse models;
Helicobacter pylori chronic infection and mucosal inflammation switches the human gastric glycosylation pathways by Ana Magalhães; Ricardo Marcos-Pinto; Alison V. Nairn; Mitche dela Rosa; Rui M. Ferreira; Susana Junqueira-Neto; Daniela Freitas; Joana Gomes; Patrícia Oliveira; Marta R. Santos; Nuno T. Marcos; Wen Xiaogang; Céu Figueiredo; Carla Oliveira; Mário Dinis-Ribeiro; Fátima Carneiro; Kelley W. Moremen; Leonor David; Celso A. Reis (1928-1939).
Helicobacter pylori exploits host glycoconjugates to colonize the gastric niche. Infection can persist for decades promoting chronic inflammation, and in a subset of individuals lesions can silently progress to cancer. This study shows that H. pylori chronic infection and gastric tissue inflammation result in a remodeling of the gastric glycophenotype with increased expression of sialyl-Lewis a/x antigens due to transcriptional up-regulation of the B3GNT5, B3GALT5, and FUT3 genes. We observed that H. pylori infected individuals present a marked gastric local pro-inflammatory signature with significantly higher TNF-α levels and demonstrated that TNF-induced activation of the NF-kappaB pathway results in B3GNT5 transcriptional up-regulation. Furthermore, we show that this gastric glycosylation shift, characterized by increased sialylation patterns, favors SabA-mediated H. pylori attachment to human inflamed gastric mucosa. This study provides novel clinically relevant insights into the regulatory mechanisms underlying H. pylori modulation of host glycosylation machinery, and phenotypic alterations crucial for life-long infection. Moreover, the biosynthetic pathways here identified as responsible for gastric mucosa increased sialylation, in response to H. pylori infection, can be exploited as drug targets for hindering bacteria adhesion and counteract the infection chronicity.
Keywords: Helicobacter pylori; Chronic infection; Glycophenotype; Sialic acid-binding adhesin;
Over-expression of neurotrophin 3 in human aortic valves affected by calcific disease induces the osteogenic responses via the Trk–Akt pathway by Qingzhou Yao; Rui Song; Lihua Ao; Qiong Zhan; Joseph C. Cleveland; Xiyong Yu; David A. Fullerton; Xianzhong Meng (1940-1949).
Calcific aortic valve disease (CAVD) is a leading cardiovascular disorder in the elderly. While aortic valve interstitial cells (AVICs) are the main cells that express osteogenic mediators, the molecular mechanism that mediates AVIC osteogenic responses is incompletely understood. This study aims to identify pro-osteogenic factors in human AVICs affected by CAVD.Microarray analysis identified 11 up-regulated genes in AVICs of diseased valves. Among these genes, mRNA levels of neurotrophin 3 (NT3) increased by 2 fold. Higher levels of NT3 protein in diseased aortic valves and diseased AVICs were confirmed by immunofluorescent staining and immunoblotting, respectively. An exposure of AVICs of normal valves to recombinant human NT3 (0.025–0.10 μg/mL) up-regulated the production of Runx2, TGF-β1 and BMP-2 in a dose-dependent fashion. NT3 also promotes calcium deposit formation. The pro-osteogenic effect of NT3 was not affected by neutralization of Toll-like receptor 2 or 4. Interestingly, mRNA encoding neural growth factor receptors (TrkA, TrkB, TrkC and p75 NTR) was detectable in human AVICs. Inhibition of Trk receptors markedly reduced the effects of NT3 on Runx2, TGF-β1 and BMP-2 production, calcium deposit formation and Akt phosphorylation. Further, inhibition of Akt also reduced the pro-osteogenic effects of NT3.AVICs of diseased human aortic valves express higher levels of NT3. NT3 up-regulates the production of Runx2, TGF-β1 and BMP-2, and promotes calcium deposit formation in human AVICs via the Trk–Akt pathway. Thus, NT3 is a novel pro-osteogenic factor in aortic valves and may play a role in valvular calcification.Display Omitted
Keywords: Neurotrophin 3; Aortic valve; Pro-osteogenic mediators; Trk; Akt;
SUMOylation of the brain-predominant Ataxin-3 isoform modulates its interaction with p97 by Bruno Almeida; Isabel A. Abreu; Carlos A. Matos; Joana S. Fraga; Sara Fernandes; Maria G. Macedo; Ricardo Gutiérrez-Gallego; Pedro José Barbosa Pereira; Ana Luísa Carvalho; Sandra Macedo-Ribeiro (1950-1959).
Machado–Joseph Disease (MJD), a form of dominantly inherited ataxia belonging to the group of polyQ expansion neurodegenerative disorders, occurs when a threshold value for the number of glutamines in Ataxin-3 (Atx3) polyglutamine region is exceeded. As a result of its modular multidomain architecture, Atx3 is known to engage in multiple macromolecular interactions, which might be unbalanced when the polyQ tract is expanded, culminating in the aggregation and formation of intracellular inclusions, a unifying fingerprint of this group of neurodegenerative disorders. Since aggregation is specific to certain brain regions, localization-dependent posttranslational modifications that differentially affect Atx3 might also contribute for MJD.We combined in vitro and cellular approaches to address SUMOylation in the brain-predominant Atx3 isoform and assessed the impact of this posttranslational modification on Atx3 self-assembly and interaction with its native partner, p97.We demonstrate that Atx3 is SUMOylated at K356 both in vitro and in cells, which contributes for decreased formation of amyloid fibrils and for increased affinity towards p97.These findings highlight the role of SUMOylation as a regulator of Atx3 function, with implications on Atx3 protein interaction network and self-assembly, with potential impact for further understanding the molecular mechanisms underlying MJD pathogenesis.
Keywords: Polyglutamine; Posttranslational modification; Protein aggregation; Amyloid; Surface plasmon resonance;
Altered protein expression pattern in skin fibroblasts from parkin-mutant early-onset Parkinson's disease patients by Rosa Lippolis; Rosa Anna Siciliano; Consiglia Pacelli; Anna Ferretta; Maria Fiorella Mazzeo; Salvatore Scacco; Francesco Papa; Antonio Gaballo; Claudia Dell'Aquila; Michele De Mari; Sergio Papa; Tiziana Cocco (1960-1970).
Parkinson's disease (PD) is the most common neurodegenerative movement disorder caused primarily by selective degeneration of the dopaminergic neurons in substantia nigra. In this work the proteomes extracted from primary fibroblasts of two unrelated, hereditary cases of PD patients, with different parkin mutations, were compared with the proteomes extracted from commercial adult normal human dermal fibroblasts (NHDF) and primary fibroblasts from the healthy mother of one of the two patients. The results show that the fibroblasts from the two different cases of parkin-mutant patients display analogous alterations in the expression level of proteins involved in different cellular functions, like cytoskeleton structure–dynamics, calcium homeostasis, oxidative stress response, protein and RNA processing.Display Omitted
Keywords: Parkinson's disease; Human skin fibroblasts; Two-dimensional gel electrophoresis; Mass spectrometry; Proteomics;
Positive feedback of DDX6/c-Myc/PTB1 regulated by miR-124 contributes to maintenance of the Warburg effect in colon cancer cells by Kohei Taniguchi; Nobuhiko Sugito; Minami Kumazaki; Haruka Shinohara; Nami Yamada; Nobuhisa Matsuhashi; Manabu Futamura; Yuko Ito; Yoshinori Otsuki; Kazuhiro Yoshida; Kazuhisa Uchiyama; Yukihiro Akao (1971-1980).
The human DEAD/H-box RNA helicase gene DDX6 is a target of the t(11;14)(q23;q32) chromosomal translocation observed in human B-cell lymphoma, and the overexpression of its protein has been shown to cause malignant transformation. DDX6 has a variety of functions such as translation initiation, pre-mRNA splicing, ribosome assembly, and more. However, details of the regulatory mechanism of DDX6 and functions of DDX6 in cancer cells are largely unknown. On the other hand, the Warburg effect is a well-known feature of cancer cells. Pyruvate kinase in muscle (PKM), which is a rate-limiting glycolytic enzyme, has 2 isoforms, PKM1 and PKM2. It has been frequently reported that PKM2 is a tumor-specific isoform and promotes the Warburg effect. However, the functions of the PKM1 gene have been hardly mentioned. Here, we showed that DDX6 was overexpressed in colorectal cancer specimens and regulated by microRNA (miR)-124 in colon cancer cells. Also, a DDX6/c-Myc/PTB1 positive feedback circuit regulated by miR-124 was shown to be established and to contribute to maintenance of the Warburg effect. Moreover, we showed that knockdown of DDX6 induced mainly apoptosis through an imbalance of PKM gene expression, especially causing down-regulation of PKM1 in colon cancer cells. These results suggest that miR-124 is a fine tuner of the Warburg effect and that DDX6 is one of the key molecules in Warburg effect-related miR-124 targeting various genes.
Keywords: DDX6; MiR-124; Warburg effect; PKM; PTBP1; c-Myc;
High-mobility group box 1 suppresses resolvin D1-induced phagocytosis via induction of resolvin D1-inactivating enzyme, 15-hydroxyprostaglandin dehydrogenase by Gyeoung-Jin Kang; Hye-Ja Lee; Yun Pyo Kang; Eun Ji Kim; Hyun Ji Kim; Hyun Jung Byun; Mi Kyung Park; Hoon Cho; Sung Won Kwon; Chang-Hoon Lee (1981-1988).
High-mobility group box 1 (HMGB1) enhances inflammatory reactions by potentiating the activity of pro-inflammatory mediators and suppressing the phagocytosis of apoptotic neutrophils. However, the effects of HMGB1 on phagocytosis induced by pro-resolving mediators, such as resolvins, have not been studied up until this point. In this study, we investigated the effects and underlying mechanism of HMGB1 on resolvin D1-induced phagocytosis of MDA-MB-231 cells, which were selected as a model system based on their phagocytic capability and ease of transfecting them with a plasmid or siRNA in several cancer cell lines. Then we confirmed effects of HMGB1 in THP-1 cells.Resolvin D1 (RvD1) enhanced phagocytosis in MDA-MB-231 and THP-1 cells. HMGB1 suppressed RvD1-induced phagocytosis in MDA-MB.231 and THP-1 cells. HMGB1 dose-dependently induced the expression of 15-hydroxyprostaglandin dehydrogenase (15-PGDH), the inactivating enzyme in pro-resolving lipid mediators such as RvE1 and RvD1. Involvement of 15-PGDH in-HMGB-1-induced suppression of phagocytosis was examined using siRNA of 15-PGDH or 15-PGDH inhibitor, TD23. Surprisingly, the silencing of 15-PGDH increased phagocytotic activity of MDA-MB-231 cells. TD23 also enhanced phagocytosis of MDA-MB-231 and THP-1 cells.In conclusion, the release of HMGB1 during the inflammatory phase induces 15-PGDH expression, which suppresses the phagocytotic activity of macrophages. These processes might be involved in the mechanism that blocks the resolution of inflammation, thereby allowing acute inflammation to progress to chronic inflammation.Display Omitted
Keywords: Resolution of inflammation; Phagocytosis; HMGB1; Resolvin D1; 15-PGDH;
Proteomics profiling of cholangiocarcinoma exosomes: A potential role of oncogenic protein transferring in cancer progression by Suman Dutta; Onrapak Reamtong; Wittaya Panvongsa; Sarunya Kitdumrongthum; Keatdamrong Janpipatkul; Polkit Sangvanich; Pawinee Piyachaturawat; Arthit Chairoungdua (1989-1999).
Cholangiocarcinoma (CCA), a common primary malignant tumor of bile duct epithelia, is highly prevalent in Asian countries and unresponsive to chemotherapeutic drugs. Thus, a newly recognized biological entity for early diagnosis and treatment is highly needed. Exosomes are small membrane bound vesicles found in body fluids and released by most cell types including cancer cells. The vesicles contain specific subset of proteins and nucleic acids corresponding to cell types and play essential roles in pathophysiological processes. The present study aimed to assess the protein profiles of CCA-derived exosomes and their potential roles. We have isolated exosomes from CCA cells namely KKU-M213 and KKU-100 derived from Thai patients and their roles were investigated by incubation with normal human cholangiocyte (H69) cells. Exosomes were internalized into H69 cells and had no effects on viability or proliferation of the host cells. Interestingly, the exosomes from KKU-M213 cells only induced migration and invasion of H69 cells. Proteomic analysis of the exosomes from KKU-M213 cells disclosed multiple cancer related proteins that are not present in H69 exosomes. Consistent with the protein profile, treatment with KKU-M213 exosomes induced β-catenin and reduced E-cadherin expressions in H69 cells. Collectively, our results suggest that a direct cell-to-cell transfer of oncogenic proteins via exosomal pathway may be a novel mechanism for CCA progression and metastasis.
Keywords: Cholangiocarcinoma; Exosomes; Proteomics; Invasion; Migration; Cell–cell communication;
Osteogenic changes in kidneys of hyperoxaluric rats by Sunil Joshi; William L. Clapp; Wei Wang; Saeed R. Khan (2000-2012).
Many calcium oxalate (CaOx) kidney stones develop attached to renal papillary sub-epithelial deposits of calcium phosphate (CaP), called Randall's plaque (RP). Pathogenesis of the plaques is not fully understood. We hypothesize that abnormal urinary environment in stone forming kidneys leads to epithelial cells losing their identity and becoming osteogenic. To test our hypothesis male rats were made hyperoxaluric by administration of hydroxy-l-proline (HLP). After 28 days, rat kidneys were extracted. We performed genome wide analyses of differentially expressed genes and determined changes consistent with dedifferentiation of epithelial cells into osteogenic phenotype. Selected molecules were further analyzed using quantitative-PCR and immunohistochemistry. Genes for runt related transcription factors (RUNX1 and 2), zinc finger protein Osterix, bone morphogenetic proteins (BMP2 and 7), bone morphogenetic protein receptor (BMPR2), collagen, osteocalcin, osteonectin, osteopontin (OPN), matrix-gla-protein (MGP), osteoprotegrin (OPG), cadherins, fibronectin (FN) and vimentin (VIM) were upregulated while those for alkaline phosphatase (ALP) and cytokeratins 10 and 18 were downregulated. In conclusion, epithelial cells of hyperoxaluric kidneys acquire a number of osteoblastic features but without CaP deposition, perhaps a result of downregulation of ALP and upregulation of OPN and MGP. Plaque formation may additionally require localized increases in calcium and phosphate and decrease in mineralization inhibitory potential.
Keywords: Runt related transcription factor; Bone morphogenetic protein; Vimentin; Matrix-gla-protein; Hyperoxaluria; Randall's plaque;