BBA - Molecular Basis of Disease (v.1863, #6)
Neurotoxic mechanisms by which the USP14 inhibitor IU1 depletes ubiquitinated proteins and Tau in rat cerebral cortical neurons: Relevance to Alzheimer's disease by Magdalena J. Kiprowska; Anna Stepanova; Dustin R. Todaro; Alexander Galkin; Arthur Haas; Scott M. Wilson; Maria E. Figueiredo-Pereira (1157-1170).
In Alzheimer's disease proteasome activity is reportedly downregulated, thus increasing it could be therapeutically beneficial. The proteasome-associated deubiquitinase USP14 disassembles polyubiquitin-chains, potentially delaying proteasome-dependent protein degradation. We assessed the protective efficacy of inhibiting or downregulating USP14 in rat and mouse (Usp14 axJ ) neuronal cultures treated with prostaglandin J2 (PGJ2). IU1 concentrations (HIU1 > 25 μM) reported by others to inhibit USP14 and be protective in non-neuronal cells, reduced PGJ2-induced Ub-protein accumulation in neurons. However, HIU1 alone or with PGJ2 is neurotoxic, induces calpain-dependent Tau cleavage, and decreases E1 ~ Ub thioester levels and 26S proteasome assembly, which are energy-dependent processes. We attribute the two latter HIU1 effects to ATP-deficits and mitochondrial Complex I inhibition, as shown herein. These HIU1 effects mimic those of mitochondrial inhibitors in general, thus supporting that ATP-depletion is a major mediator of HIU1-actions. In contrast, low IU1 concentrations (LIU1 ≤ 25 μM) or USP14 knockdown by siRNA in rat cortical cultures or loss of USP14 in cortical cultures from ataxia (Usp14 axJ ) mice, failed to prevent PGJ2-induced Ub-protein accumulation. PGJ2 alone induces Ub-protein accumulation and decreases E1 ~ Ub thioester levels. This seemingly paradoxical result may be attributed to PGJ2 inhibiting some deubiquitinases (such as UCH-L1 but not USP14), thus triggering Ub-protein stabilization. Overall, IU1-concentrations that reduce PGJ2-induced accumulation of Ub-proteins are neurotoxic, trigger calpain-mediated Tau cleavage, lower ATP, E1 ~ Ub thioester and E1 protein levels, and reduce proteasome activity. In conclusion, pharmacologically inhibiting (with low or high IU1 concentrations) or genetically down-regulating USP14 fail to enhance proteasomal degradation of Ub-proteins or Tau in neurons.Display Omitted
Keywords: Alzheimer's; Deubiquitinase; Ubiquitin-activating enzyme; USP14; Mitochondria; Tau; Calpain;
Translational effects and coding potential of an upstream open reading frame associated with DOPA Responsive Dystonia by Lataisia Jones; Lacy Goode; Eduardo Davila; Amber Brown; Deirdre M. McCarthy; Nutan Sharma; Pradeep G. Bhide; Ioanna A. Armata (1171-1182).
Upstream open reading frames (uORFs) have emerged as major post-transcriptional regulatory elements in eukaryotic species. In general, uORFs are initiated by a translation start codon within the 5′ untranslated region of a gene (upstream ATG; uATG), and they are negatively correlated with translational efficiency. In addition to their translational regulatory role, some uORFs can code for biologically active short peptides. The importance of uATGs/uORFs is further underscored by human diseases associated with single nucleotide polymorphisms (SNPs), which disrupt existing uORFs or introduce novel uORFs. Although several functional proteins translated from naturally occurring uORFs have been described, the coding potential of uORFs created by SNPs has been ignored because of the a priori assumption that these proteins are short-lived with no likely impact on protein homeostasis. Thus, studies on SNP-created uORFs are limited to their translational effects, leaving unexplored the potential cellular consequences of a SNP/uORF-encoded protein. Here, we investigate functionality of a uATG/uORF introduced by a + 142C > T SNP within the GCH1 gene and associated with a familial form of DOPA Responsive Dystonia. We report that the + 142C > T SNP represses GCH1 translation, and introduces a short, frame shifted uORF that encodes a 73-amino acid peptide. This peptide is localized within the nucleus and compromises cell viability upon proteasome inhibition. Our work extends the list of uATG/uORF associated diseases and advances research on peptides translated from SNP-introduced uORFs, a neglected component of the proteome.Display Omitted
Keywords: DOPA Responsive Dystonia; DYT5; Upstream open reading frames; 5′Untranslated region; Regulation of translation; Gene expression; Coding potential; Short peptides;
P2X7 receptor promotes intestinal inflammation in chemically induced colitis and triggers death of mucosal regulatory T cells by Vanessa R. Figliuolo; Luiz Eduardo Baggio Savio; Hanaa Safya; Hayandra Nanini; Cláudio Bernardazzi; Alessandra Abalo; Heitor S.P. de Souza; Jean Kanellopoulos; Pierre Bobé; Cláudia M.L.M. Coutinho; Robson Coutinho-Silva (1183-1194).
P2X7 receptor activation contributes to inflammation development in different pathologies. We previously reported that the P2X7 receptor is over-expressed in the gut mucosa of patients with inflammatory bowel disease, and that P2X7 inhibition protects against chemically induced colitis. Here, we investigated in detail the role of the P2X7 receptor in inflammatory bowel disease development, by treating P2X7 knockout (KO) and WT mice with two different (and established) colitis inductors. P2X7 KO mice were protected against gut inflammation induced by 2,4,6-trinitrobenzenesulfonic acid or oxazolone, with no weight loss or gut histological alterations after treatment. P2X7 receptor knockout induced regulatory T cell accumulation in the colon, as evaluated by qRT-PCR for FoxP3 expression and immunostaining for CD90/CD45RBlow. Flow cytometry analysis of mesenteric lymph node cells showed that P2X7 activation (by ATP) triggered regulatory T cell death. In addition, such cells from P2X7 KO mice expressed more CD103, suggesting increased migration of regulatory T cells to the colon (relative to the WT). Our results show that the P2X7 has a key role during inflammation development in inflammatory bowel disease, by triggering the death and retention in the mesenteric lymph nodes of regulatory T cells that would otherwise promote immune system tolerance in the gut.
Keywords: ATP; P2X7 receptor; Colitis, regulatory T cells;
Astragaloside IV attenuates lead acetate-induced inhibition of neurite outgrowth through activation of Akt-dependent Nrf2 pathway in vitro by Chunlei Yu; Siwen Pan; Miaoxian Dong; Yingcai Niu (1195-1203).
Recently, oxidative stress is strongly associated with lead (Pb)-induced neurotoxicity. We reported previously that Astragaloside IV (AS-IV) possesses potent antioxidant properties. Here, we evaluate the hypothesis that AS-IV attenuates lead acetate (PbAc)-mediated inhibition of neurite outgrowth might mainly result from its antioxidant property via serine/threonine protein kinase (Akt)-dependent activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. Interestingly, AS-IV attenuates PbAc-induced inhibition of neurite outgrowth and displayed potential antioxidant properties by inhibiting reactive oxygen species (ROS). Concomitantly, AS-IV enhanced phase II detoxifying enzymes such as heme oxygenase 1 (HO-1), thioredoxin reductase (TrxR), and glutamate cysteine ligase catalytic subunit (GCLc). Conversely, AS-IV had no effect on GCL modulatory subunit (GCLm) and superoxide dismutase (SOD) activity/expression. Furthermore, AS-IV evoked Akt phosphorylation, and subsequent induced phosphorylation of glycogen synthase kinase-3β (GSK-3β) at Ser9 (that is, inactivation), which stimulated Nrf2-mediated antioxidant response element (ARE)-containing activation. Importantly, Akt locates upstream of GSK-3β and regulates phase II detoxifying enzymes gene expression through Nrf2 nuclear accumulation in PC12 cells exposed to PbAc. Noteworthy, these results were further confirmed through signalling pathway inhibitors, dominant negative mutant and short hairpin RNA technology. Collectively, these in vitro findings suggest that AS-IV attenuates PbAc-induced inhibition of neurite outgrowth attributed to its antioxidant properties and may be a promising candidate for the treatment of lead developmental neurotoxicity.Display Omitted
Keywords: Astragaloside IV; Oxidative stress; Neurite outgrowth; Lead acetate; Nrf2;
7,8-Dihydroxyflavone facilitates the action exercise to restore plasticity and functionality: Implications for early brain trauma recovery by Gokul Krishna; Rahul Agrawal; Yumei Zhuang; Zhe Ying; Afshin Paydar; Neil G Harris; Luiz Fernando F. Royes; Fernando Gomez-Pinilla (1204-1213).
Metabolic dysfunction accompanying traumatic brain injury (TBI) severely impairs the ability of injured neurons to comply with functional demands. This limits the success of rehabilitative strategies by compromising brain plasticity and function, and highlights the need for early interventions to promote energy homeostasis. We sought to examine whether the TrkB agonist, 7,8-dihydroxyflavone (7,8-DHF) normalizes brain energy deficits and reestablishes more normal patterns of functional connectivity, while enhancing the effects of exercise during post-TBI period. Moderate fluid percussion injury (FPI) was performed and 7,8-DHF (5 mg/kg, i.p.) was administered in animals subjected to FPI that either had access to voluntary wheel running for 7 days after injury or were sedentary. Compared to sham-injured controls, TBI resulted in reduced hippocampal activation of the BDNF receptor TrkB and associated CREB, reduced levels of plasticity markers GAP-43 and Syn I, as well as impaired memory as indicated by the Barnes maze task. While 7,8-DHF treatment and exercise individually mitigated TBI-induced effects, administration of 7,8-DHF concurrently with exercise facilitated memory performance and augmented levels of markers of cell energy metabolism viz., PGC-1α, COII and AMPK. In parallel to these findings, resting-state functional MRI (fMRI) acquired at 2 weeks after injury showed that 7,8-DHF with exercise enhanced hippocampal functional connectivity, and suggests 7,8-DHF and exercise to promote increases in functional connectivity. Together, these findings indicate that post-injury 7,8-DHF treatment promotes enhanced levels of cell metabolism, synaptic plasticity in combination with exercise increases in brain circuit function that facilitates greater physical rehabilitation after TBI.
Keywords: 7,8-Dihydroxyflavone; Exercise; Memory; Traumatic brain injury; Functional connectivity; Rehabilitation;
Defects in mitochondrial energetic function compels Fanconi Anaemia cells to glycolytic metabolism by Enrico Cappelli; Paola Cuccarolo; Giorgia Stroppiana; Maurizio Miano; Roberta Bottega; Vanessa Cossu; Paolo Degan; Silvia Ravera (1214-1221).
Energetic metabolism plays an essential role in the differentiation of haematopoietic stem cells (HSC). In Fanconi Anaemia (FA), DNA damage is accumulated during HSC differentiation, an event that is likely associated with bone marrow failure (BMF). One of the sources of the DNA damage is altered mitochondrial metabolism and an associated increment of oxidative stress. Recently, altered mitochondrial morphology and a deficit in the energetic activity in FA cells have been reported. Considering that mitochondria are the principal site of aerobic ATP production, we investigated FA metabolism in order to understand what pathways are able to compensate for this energy deficiency. In this work, we report that the impairment in mitochondrial oxidative phosphorylation (OXPHOS) in FA cells is countered by an increase in glycolytic flux. By contrast, glutaminolysis appears lower with respect to controls. Therefore, it is possible to conclude that in FA cells glycolysis represents the main pathway for producing energy, balancing the NADH/NAD+ ratio by the conversion of pyruvate to lactate. Finally, we show that a forced switch from glycolytic to OXPHOS metabolism increases FA cell oxidative stress. This could be the cause of the impoverishment in bone marrow HSC during exit from the homeostatic quiescent state. This is the first work that systematically explores FA energy metabolism, highlighting its flaws, and discusses the possible relationships between these defects and BMF.
Keywords: Fanconi Anaemia; Energy metabolism; Glycolysis; Oxidative phosphorylation; Oxidative stress; Cancer-prone diseases;
Specific patterns of spinal metabolites underlying α-Me-5-HT-evoked pruritus compared with histamine and capsaicin assessed by proton nuclear magnetic resonance spectroscopy by Taotao Liu; Zhigang He; Xuebi Tian; Ghulam Mustafa Kamal; Zhixiao Li; Zeyuan Liu; Huili Liu; Fuqiang Xu; Jie Wang; Hongbing Xiang (1222-1230).
The mechanism behind itching is not well understood. Proton nuclear magnetic resonance (1H–NMR) spectroscopic analysis of spinal cord extracts provides a quick modality for evaluating the specific metabolic activity of α-Me-5-HT-evoked pruritus mice. In the current study, four groups of young adult male C57Bl/6 mice were investigated; one group treated with saline, while the other groups intradermally injected with α-Me-5-HT (histamine independent pruritogen), histamine (histamine dependent pruritogen) and capsaicin (algogenic substance), respectively. The intradermal microinjection of α-Me-5-HT and histamine resulted in a dramatic increase in the itch behavior. Furthermore, the results of NMR studies of the spinal cord extracts revealed that the metabolites show very different patterns for these different drugs, especially when comparing α-Me-5-HT and capsaicin. All the animals in the groups of α-Me-5-HT and capsaicin were completely separated using the metabolite parameters and principal component analysis. For α-Me-5-HT, the concentrations of glutamate, GABA, glycine and aspartate increased significantly, especially for GABA (increased 17.2%, p = 0.008). Furthermore, the concentration of NAA increased, but there was no significant difference (increased 11.3%, p = 0.191) compared to capsaicin (decreased 29.1%, p = 0.002). Thus the application of magnetic resonance spectroscopy technique, coupled with statistical analysis, could further explain the mechanism behind itching evoked by α-Me-5-HT or other drugs. It can thus improve our understanding of itch pathophysiology and pharmacological therapies which may contribute to itch relief.Display Omitted
Keywords: Itch; Spinal cord; Metabolites; Nuclear magnetic resonance spectroscopy; GABA;
Aquaporin-3 deletion in mice results in renal collecting duct abnormalities and worsens ischemia-reperfusion injury by Lei Lei; Weiling Wang; Yingli Jia; Limin Su; Hong Zhou; Alan S. Verkman; Baoxue Yang (1231-1241).
Aquaporin-3 (AQP3), a transporter of water, glycerol and H2O2, is expressed in basolateral membranes of principal cells in kidney collecting duct. Here, we report that AQP3 deletion in mice affects renal function and modulates renal injury. We found collecting duct hyperplasia and cell swelling in kidneys of adult AQP3 null mice. After mild renal ischemia-reperfusion (IR), AQP3 null mice had significantly greater blood urea nitrogen (57 mg/dl) and creatinine (136 μM) than wild-type mice (35 mg/dl and 48 μM, respectively), and showed renal morphological changes, including tubular dilatation, erythrocyte diapedesis and collecting duct incompletion. MPO, MDA and SOD following IR in AQP3 null mice were significantly different from that in wild-type mice (1.7 U/g vs 0.8 U/g, 3.9 μM/g vs 2.4 μM/g, 6.4 U/mg vs 11 U/mg, respectively). Following IR, AQP3 deletion inhibited activation of mitogen-activated protein kinase (MAPK) signaling and produced an increase in the ratios of Bax/Bcl-2, cleaved caspase-3/caspase-3 and p-p53/p53. Studies in transfected MDCK cells showed that AQP3 expression attenuated reduced cell viability following hypoxia-reoxygenation, with reduced apoptosis and increased MAPK signaling. Our results support a novel role for AQP3 in modulating renal injury and suggest the mechanisms involved in protection against hypoxic injury.
Keywords: AQP3; Ischemic injury; Acute kidney injury; MDCK; Kidney; MAPK;
Impaired vitreous composition and retinal pigment epithelium function in the FoxG1::LRP2 myopic mice by Olivier Cases; Antoine Obry; Sirine Ben-Yacoub; Sébastien Augustin; Antoine Joseph; Géraldine Toutirais; Manuel Simonutti; Annabel Christ; Pascal Cosette; Renata Kozyraki (1242-1254).
High myopia (HM) is one of the main causes of visual impairment and blindness all over the world and an unsolved medical problem. Persons with HM are predisposed to other eye pathologies such as retinal detachment, myopic retinopathy or glaucomatous optic neuropathy, complications that may at least partly result from the extensive liquefaction of the myopic vitreous gel. To identify the involvement of the liquid vitreous in the pathogenesis of HM we here analyzed the vitreous of the recently described highly myopic low density lipoprotein receptor-related protein 2 (Lrp2)-deficient eyes. Whereas the gel-like fraction was not apparently modified, the volume of the liquid vitreous fraction (LVF) was much higher in the myopic eyes. Biochemical and proteome analysis of the LVF revealed several modifications including a marked decrease of potassium, sodium and chloride, of proteins involved in ocular tissue homeostasis and repair as well as of ADP-ribosylation factor 4 (ARF4), a protein possibly involved in LRP2 trafficking. A small number of proteins, mainly comprising known LRP2 ligands or proteins of the inflammatory response, were over expressed in the mutants. Moreover the morphology of the LRP2-deficient retinal pigment epithelium (RPE) cells was affected and the expression of ARF4 as well as of proteins involved in degradative endocytosis was strongly reduced. Our results support the idea that impairment of the RPE structure and most likely endocytic function may contribute to the vitreal modifications and pathogenesis of HM.
Keywords: High myopia; Endocytosis; Vitreous; low density lipoprotein receptor-related protein 2;
High-sucrose-induced maternal obesity disrupts ovarian function and decreases fertility in Drosophila melanogaster by Rita T. Brookheart; Alison R. Swearingen; Christina A. Collins; Laura M. Cline; Jennifer G. Duncan (1255-1263).
As the obesity epidemic worsens, the prevalence of maternal obesity is expected to rise. Both high-fat and high-sucrose diets are known to promote maternal obesity and several studies have elucidated the molecular influence of high-fat feeding on female reproduction. However, to date, the molecular impact of a high-sucrose diet on maternal obesity remains to be investigated. Using our previously reported Drosophila high-sucrose maternal obesity model, we sought to determine how excess dietary sucrose impacted the ovary. High-sucrose diet (HSD) fed adult females developed systemic insulin resistance and exhibited an ovarian phenotype characterized by excess accumulation of lipids and cholesterol in the ovary, decreased ovary size, and impaired egg maturation. We also observed decreased expression of antioxidant genes and increased protein carbonylation in the ovaries of HSD females. HSD females laid fewer eggs; however, the overall survival of offspring was unchanged relative to lean control females. Ovaries of HSD females had increased mitochondrial DNA copy number and decreased expression of key mitochondrial regulators, suggestive of an ineffective compensatory response to mitochondrial dysfunction. Mitochondrial alterations were also observed in male offspring of obese females. This study demonstrates that high-sucrose-induced maternal obesity promotes insulin resistance, while disrupting ovarian metabolism and function.
Keywords: Obesity; Drosophila; Ovary; Mitochondria; Pregnancy; Nutritional programming;
Relationship of angiotensin I-converting enzyme (ACE) and bradykinin B2 receptor (BDKRB2) polymorphism with diabetic nephropathy by Honghong Zou; Guoqing Wu; Jinlei Lv; Gaosi Xu (1264-1272).
We provide first evidence indicating the causation between ACE DD or B2R + 9 bp genotype and the increased risk for diabetic nephropathy, broadening our horizon about the role of genetic modulators in this disease.
Keywords: ACE I/D polymorphism; BDKRB2 + 9/− 9 polymorphism; Diabetic nephropathy; Mitochondrial oxidative stress;
NCLs and ER: A stressful relationship by Davide Marotta; Elisa Tinelli; Sara E. Mole (1273-1281).
The Neuronal Ceroid Lipofuscinoses (NCLs, Batten disease) are a group of inherited neurodegenerative disorders with variable age of onset, characterized by the lysosomal accumulation of autofluorescent ceroid lipopigments. The endoplasmic reticulum (ER) is a critical organelle for normal cell function. Alteration of ER homeostasis leads to accumulation of misfolded protein in the ER and to activation of the unfolded protein response. ER stress and the UPR have recently been linked to the NCLs. In this review, we will discuss the evidence for UPR activation in the NCLs, and address its connection to disease pathogenesis. Further understanding of ER-stress response involvement in the NCLs may encourage development of novel therapeutical agents targeting these pathogenic pathways.
Keywords: Batten disease; CLN1; CLN3; CLN6; CLN8; ER stress;
Dihydromyricetin prevents obesity-induced slow-twitch-fiber reduction partially via FLCN/FNIP1/AMPK pathway by Qicheng Zhou; Yeyun Gu; Hedong Lang; Xiaolan Wang; Ka Chen; Xinhua Gong; Min Zhou; Li Ran; Jundong Zhu; Mantian Mi (1282-1291).
Obesity is often accompanied by decreases in the proportion of skeletal muscle slow-twitch fibers and insulin sensitivity. Increased plasma non-esterified fatty acids (NEFA) levels are responsible for obesity-associated insulin resistance. Palmitate, one of the most elevated plasma NEFA in obesity, has been recognized as the principle inducer of insulin resistance. The present study showed that increased plasma NEFA levels were negatively linked to slow-twitch fiber proportion and insulin sensitivity, while slow-twitch fiber proportion was positively correlated to insulin sensitivity in high fat diet (HFD)-fed and ob/ob mice. Dihydromyricetin (DHM) intervention increased slow-twitch fiber proportion and improved insulin resistance. In cultured C2C12 myotubes, palmitate treatment resulted in decrease of slow-twitch fiber specific Myh7 expression and insulin resistance, concomitant with folliculin (FLCN) and folliculin-interacting protein 1 (FNIP1) expression increase, AMP-activated protein kinase (AMPK) inactivation and peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) expression decrease. Those palmitate-induced effects could be blocked by knock-down of FLCN expression or DHM intervention. Meanwhile, the protective effects of DHM were alleviated by over-expression of FLCN. In addition, the changes in AMPK activity and expression of FLCN and FNIP1 in vivo were consistent with those occurring in vitro. These findings suggest that DHM treatment prevents palmitate-induced slow-twitch fibers decrease partially via FLCN-FNIP1-AMPK pathway thereby improving insulin resistance in obesity.
Keywords: AMPK; Dihydromyricetin; FLCN; FNIP1; Slow fibers;
Pharmacological inhibition of carnitine palmitoyltransferase 1 restores mitochondrial oxidative phosphorylation in human trifunctional protein deficient fibroblasts by Bruno Lefort; Elodie Gouache; Cécile Acquaviva; Marine Tardieu; Jean François Benoist; Jean-François Dumas; Stéphane Servais; Stéphan Chevalier; Christine Vianey-Saban; François Labarthe (1292-1299).
Mitochondrial Trifunctional Protein deficiency (TFPD) is a severe genetic disease characterized by altered energy metabolism and accumulation of long-chain (LC) acylcarnitines in blood and tissues. This accumulation could impair the mitochondrial oxidative phosphorylation (OxPhos), contributing to the non-optimal outcome despite conventional diet therapy with medium-chain triglycerides (MCT).Acylcarnitine and OxPhos parameters were measured in TFPD-fibroblasts obtained from 8 children and cultured in medium mimicking fasting (LCFA) or conventional treatment (MCT), with or without Etomoxir (ETX) an inhibitor of carnitine palmitoyltransferase 1 (CPT1) activity, and were compared to results obtained with fibroblasts from 5 healthy-control children. The effects of various acylcarnitines were also tested on control fibroblasts.In the LCFA-condition, TFPD-fibroblasts demonstrated a large accumulation of LC-acylcarnitines associated with decreased O2-consumption (63 ± 3% of control, P < 0.001) and ATP production (67 ± 5%, P < 0.001) without modification of coupling efficiency. A dose-dependent decrease in O2-consumption was reproduced in control fibroblasts by addition of increasing dose of LC-acylcarnitines, while it was almost preserved with MC-acylcarnitines. The MCT-condition reduced LC-acylcarnitine accumulation and partially improved O2-consumption (80 ± 3%, P < 0.01) in TFPD-fibroblasts. The addition of ETX in both LCFA- and MCT-conditions normalized acylcarnitine profiles and restored O2-consumption and ATP production at the same levels than control.Accumulation of LC-acylcarnitines plays a major role in the pathophysiology of TFPD, reducing OxPhos capacities. These deleterious effects could be partially prevented by MCT-therapy and totally corrected by ETX. Inhibition of CPT1 may be view as a new therapeutic target for patients with a severe form of TFPD.
Keywords: Mitochondrial trifunctional protein deficiency; Long-chain 3-hydroxyacyl-CoA dehydrogenase; Mitochondria; Acylcarnitines; Oxidative phosphorylation; Etomoxir;
Nogo-A in the visual system development and in ocular diseases by Vincent Pernet (1300-1311).
Nogo-A is a potent myelin-associated inhibitor for neuronal growth and plasticity in the central nervous system (CNS). Its effects are mediated by the activation of specific receptors that intracellularly control cytoskeleton rearrangements, protein synthesis and gene expression. Moreover, Nogo-A has been involved in the development of the visual system and in a variety of neurodegenerative diseases and injury processes that can alter its function. For example, Nogo-A was shown to influence optic nerve myelinogenesis, the formation and maturation of retinal axon projections, and retinal angiogenesis. In adult animals, the inactivation of Nogo-A exerted remarkable effects on visual plasticity. Relieving Nogo-A-induced inhibition increased axonal sprouting after optic nerve lesion and axonal rewiring in the visual cortex of intact adult mice. This review aims at presenting our current knowledge on the role of Nogo-A in the visual system and to discuss how its therapeutic targeting may promote visual improvement in ophthalmic diseases.
Keywords: Nogo-A; Myelinogenesis; Retinal ganglion cells; Axonal regeneration; Multiple sclerosis; Amyotrophic lateral sclerosis;
Insulin increases filtration barrier permeability via TRPC6-dependent activation of PKGIα signaling pathways by Dorota Rogacka; Irena Audzeyenka; Patrycja Rachubik; Michał Rychłowski; Małgorzata Kasztan; Maciej Jankowski; Stefan Angielski; Agnieszka Piwkowska (1312-1325).
Podocytes are dynamic polarized cells on the surface of glomerular capillaries and an essential component of the glomerular filtration barrier. Insulin increases the activation of protein kinase G type Iα (PKGIα) subunits, leading to podocyte dysfunction. In addition, accumulating evidence suggests that TRPC6 channels are crucial mediators of podocyte calcium handling and involved in the regulation of glomerular filtration. Therefore, we investigated whether TRPC6 is involved in the regulation of filtration barrier permeability by insulin via the PKGIα-dependent manner.TRPC channel inhibitor SKF96365 abolished insulin-dependent glomerular albumin permeability and transepithelial albumin flux in cultured rat podocytes. Insulin-evoked albumin permeability across podocyte monolayers was also blocked using TRPC6 siRNA. The effect of insulin on albumin permeability was mimicked by treating podocytes with TRPC channel activator (oleolyl-2-acetyl-sn-glycerol, OAG). Insulin or OAG treatment rapidly increased the superoxide generation through activation of NADH oxidase. TRPC inhibitor SKF96365 or siRNA knockdown of TRPC6 attenuated insulin-dependent increase of ROS production. Furthermore, TRPC inhibitor or downregulation of TRPC6 blocked insulin-induced rearrangement of the actin cytoskeleton and attenuated oxidative activation of PKGIα and changes in the phosphorylation of PKG target proteins MYPT1 and MLC. Moreover insulin regulated the PKGIα interaction with TRPC6 in cultured rat podocytes.Taken together, our data suggest a key role of TRPC6 channels in the mediation of insulin-dependent activation of PKGIα signaling pathways. Overall, we have identified a potentially important mechanism that may explain disturbances in filtration barrier permeability in many diseases with increased expression of TRPC6 and chronic Ca2 + overload.
Keywords: Insulin; Filtration barrier permeability; Podocyte; Protein kinase G type I alpha; TRPC6; NADPH oxidase;
Endothelial cell calpain as a critical modulator of angiogenesis by Yixuan Zhang; Norika Mengchia Liu; Yongchen Wang; Ji Youn Youn; Hua Cai (1326-1335).
Calpains are a family of calcium-dependent non-lysosomal cysteine proteases. In particular, calpains residing in the endothelial cells play important roles in angiogenesis. It has been shown that calpain activity can be increased in endothelial cells by growth factors, primarily vascular endothelial growth factor (VEGF). VEGF/VEGFR2 induces calpain 2 dependent activation of PI3K/AMPK/Akt/eNOS pathway, and consequent nitric oxide production and physiological angiogenesis. Under pathological conditions such as tumor angiogenesis, endothelial calpains can be activated by hypoxia. This review focuses on the molecular regulatory mechanisms of calpain activation, and the newly identified mechanistic roles and downstream signaling events of calpains in physiological angiogenesis, and in the conditions of pathological tumor angiogenesis and diabetic wound healing, as well as retinopathy and atherosclerosis that are also associated with an increase in calpain activity. Further discussed include the differential strategies of modulating angiogenesis through manipulating calpain expression/activity in different pathological settings. Targeted limitation of angiogenesis in cancer and targeted promotion of angiogenesis in diabetic wound healing via modulations of calpains and calpain-dependent signaling mechanisms are of significant translational potential. Emerging strategies of tissue-specific targeting, environment-dependent targeting, and genome-targeted editing may turn out to be effective regimens for targeted manipulation of angiogenesis through calpain pathways, for differential treatments including both attenuation of tumor angiogenesis and potentiation of diabetic angiogenesis.
Keywords: Angiogenesis; Calpain; Endothelial cells; VEGF; eNOS; Tumor angiogenesis; Diabetic wound healing; Atherosclerosis; Retinopathy; Shear stress;
Dual role of MUC1 mucin in kidney ischemia-reperfusion injury: Nephroprotector in early phase, but pro-fibrotic in late phase by Jean-Baptiste Gibier; Brigitte Hémon; Mélanie Fanchon; Kelly Gaudelot; Nicolas Pottier; Bélinda Ringot; Isabelle Van Seuningen; François Glowacki; Christelle Cauffiez; David Blum; Marie-Christine Copin; Michaël Perrais; Viviane Gnemmi (1336-1349).
Acute kidney injury (AKI) is characterized by acute tubular necrosis (ATN) which involves mainly proximal tubules. Past AKI is associated with higher risk of chronic kidney disease (CKD). The MUC1 mucin is a large glycoprotein responsible for epithelial protection and locates to convoluted distal tubules and collecting ducts. Since MUC1 activates the epithelial-mesenchymal transition (EMT) in carcinoma cells, we hypothesized that MUC1 could be involved in epithelial tubular cell plasticity, a process that not only accompanies epithelial repair, but also participates into kidney fibrosis, histological substratum of CKD. In cultured human proximal cells and in human kidney allograft biopsies, we observed MUC1 induction in proximal tubules displaying ATN. Transient MUC1 induction localized with mesenchymal and stem-cell markers and was associated in vitro with reduced anoikis. In a mouse ischemia-reperfusion (IR) model, Muc1 expression mitigates severe tubular injury, as WT displayed less ATN than Muc1 KO mice. But, WT mice displayed more severe kidney fibrosis than Muc1 KO 28 days after ischemia. Besides, sustained Muc1 expression in WT was associated with less kidney M2 macrophages. Human kidney biopsies performed within the first week (W1) of transplantation in the context of IR showed MUC1 W1 induction associated with EMT markers. Protocol biopsies performed 3 months after demonstrated sustained abnormal MUC1 induction in atrophic tubules within kidney fibrosis. Altogether these data showed that sustained abnormal MUC1 induction accompanies failing epithelial repair, chronic inflammation and kidney fibrosis. In conclusion, MUC1 exerts opposite effects during kidney response to IR: first protective and then harmful.
Keywords: MUC1; Acute kidney injury; Epithelial-mesenchymal transition; Ischemia-reperfusion; Fibrosis;
Annexin A1 nuclear translocation induces retinal ganglion cell apoptosis after ischemia-reperfusion injury through the p65/IL-1β pathway by Yin Zhao; Xing Li; Jieling Gong; Lu Li; Liwen Chen; Lu Zheng; Zhiqi Chen; Jing Shi; Hong Zhang (1350-1358).
The degeneration of retinal ganglion cells (RGCs) has been identified as a major problem in glaucoma. Previous studies have indicated an association between annexin A1 (ANXA1) and neuronal cell apoptosis, and RGCs apoptosis in acute ischemia-reperfusion was attributed to an increased production of IL-1β. We found that the expression and nuclear translocation of ANXA1 were upregulated in models of acute ischemia-reperfusion in RGCs in vivo. ANXA1 was found to have a promoting effect on the expression of IL-1β in primary cultured RGCs, which could be inhibited by treatment with ANXA1 shRNA or the p65 inhibitor BAY 11-7082. ANXA1 interacted with p65, and recruited it into the nucleus. Chromatin immunoprecipitation assay revealed that ANXA1 accumulated at the IL-1β gene promoter. The reduction of p65 nuclear translocation using a membrane-permeable ANXA1 peptide containing a Ser5Ala mutation led to a decrease in the expression of IL-1β, and acute ischemia-reperfusion induced RGCs apoptosis in vivo. These results indicate that in RGCs, ANXA1 increases IL-1β expression by recruiting p65 to the nucleus, which induces cell apoptosis. The obtained results may help the development of a novel treatment strategy against RGCs apoptosis in acute ischemia-reperfusion injury.
Keywords: Annexin A1; Nuclear translocation; Retinal ganglion cell; IL-1β; Apoptosis;
Mfn2 protects dopaminergic neurons exposed to paraquat both in vitro and in vivo: Implications for idiopathic Parkinson's disease by Fanpeng Zhao; Wenzhang Wang; Chunyu Wang; Sandra L. Siedlak; Hisashi Fujioka; Beisha Tang; Xiongwei Zhu (1359-1370).
Mitochondrial dynamics and quality control play a critical role in the maintenance of mitochondrial homeostasis and function. Pathogenic mutations of many genes associated with familial Parkinson's disease (PD) caused abnormal mitochondrial dynamics, suggesting a likely involvement of disturbed mitochondrial fission/fusion in the pathogenesis of PD. In this study, we focused on the potential role of mitochondrial fission/fusion in idiopathic PD patients and in neuronal cells and animals exposed to paraquat (PQ), a commonly used herbicide and PD-related neurotoxin, as models for idiopathic PD. Significantly increased expression of dynamin-like protein 1 (DLP1) and a trend towards reduced expression of Mfn1 and Mfn2 were noted in the substantia nigra tissues from idiopathic PD cases. Interestingly, PQ treatment led to similar changes in the expression of fission/fusion proteins both in vitro and in vivo which was accompanied by extensive mitochondrial fragmentation and mitochondrial dysfunction. Blockage of PQ-induced mitochondrial fragmentation by Mfn2 overexpression protected neurons against PQ-induced mitochondrial dysfunction in vitro. More importantly, PQ-induced oxidative damage and stress signaling as well as selective loss of dopaminergic (DA) neurons in the substantia nigra and axonal terminals in striatum was also inhibited in transgenic mice overexpressing hMfn2. Overall, our study demonstrated that disturbed mitochondrial dynamics mediates PQ-induced mitochondrial dysfunction and neurotoxicity both in vitro and in vivo and is also likely involved in the pathogenesis of idiopathic PD which make them a promising therapeutic target for PD treatment.
Keywords: Mitochondrial dynamics; DLP1; Drp1; Mfn2; Paraquat; Parkinson's disease;
Paternal low protein diet programs preimplantation embryo gene expression, fetal growth and skeletal development in mice by Adam J. Watkins; Slobodan Sirovica; Ben Stokes; Mark Isaacs; Owen Addison; Richard A. Martin (1371-1381).
Defining the mechanisms underlying the programming of early life growth is fundamental for improving adult health and wellbeing. While the association between maternal diet, offspring growth and adult disease risk is well-established, the effect of father's diet on offspring development is largely unknown. Therefore, we fed male mice an imbalanced low protein diet (LPD) to determine the impact on post-fertilisation development and fetal growth. We observed that in preimplantation embryos derived from LPD fed males, expression of multiple genes within the central metabolic AMPK pathway was reduced. In late gestation, paternal LPD programmed increased fetal weight, however, placental weight was reduced, resulting in an elevated fetal:placental weight ratio. Analysis of gene expression patterns revealed increased levels of transporters for calcium, amino acids and glucose within LPD placentas. Furthermore, placental expression of the epigenetic regulators Dnmt1 and Dnmt3L were increased also, coinciding with altered patterns of maternal and paternal imprinted genes. More strikingly, we observed fetal skeletal development was perturbed in response to paternal LPD. Here, while offspring of LPD fed males possessed larger skeletons, their bones comprised lower volumes of high mineral density in combination with reduced maturity of bone apatite. These data offer new insight in the underlying programming mechanisms linking poor paternal diet at the time of conception with the development and growth of his offspring.
Keywords: Blastocyst metabolism; Bone health; Developmental programming; Fetal growth; Paternal diet; Placental function;
Endothelial Nox4-based NADPH oxidase regulates atherosclerosis via soluble epoxide hydrolase by Pingping Hu; Xiaojuan Wu; Alok R. Khandelwal; Weimin Yu; Zaicheng Xu; Lili Chen; Jian Yang; Robert M. Weisbrod; Kin Sing Stephen Lee; Francesca Seta; Bruce D. Hammock; Richard A. Cohen; Chunyu Zeng; Xiaoyong Tong (1382-1391).
Nox4-based NADPH oxidase is a major reactive oxygen species-generating enzyme in the vasculature, but its role in atherosclerosis remains controversial.Our goal was to investigate the mechanisms of endothelial Nox4 in regulating atherosclerosis.Atherosclerosis-prone conditions (disturbed blood flow, type I diabetes, and Western diet) downregulated endothelial Nox4 mRNA in arteries. To address whether the downregulated endothelial Nox4 was directly involved in the development of atherosclerosis, we generated mice carrying a human Nox4 P437H dominant negative mutation (Nox4DN), driven by the endothelial specific promoter Tie-2, on atherosclerosis-prone genetic background (ApoE deficient mice) to mimic the effect of decreased endothelial Nox4. Nox4DN significantly increased type I diabetes-induced aortic stiffness and atherosclerotic lesions. Gene analysis indicated that soluble epoxide hydrolase 2 (sEH) was significantly upregulated in Nox4DN endothelial cells (EC). Inhibition of sEH activity in Nox4DN EC suppressed inflammation and macrophage adhesion to EC. On the contrary, overexpression of endothelial wild type Nox4 suppressed sEH, ameliorated Western diet-induced atherosclerosis and decreased aortic stiffness.Atherosclerosis-prone conditions downregulated endothelial Nox4 to accelerate the progress of atherosclerosis, at least in part, by upregulating sEH to enhance inflammation.
Keywords: Nox4; Atherosclerosis; Endothelium; Soluble epoxide hydrolase 2; Type I diabetes;
Assessment of histone tail modifications and transcriptional profiling during colon cancer progression reveals a global decrease in H3K4me3 activity by Karen Triff; Mathew W. McLean; Kranti Konganti; Jiahui Pang; Evelyn Callaway; Beiyan Zhou; Ivan Ivanov; Robert S. Chapkin (1392-1402).
During colon cancer, epigenetic alterations contribute to the dysregulation of major cellular functions and signaling pathways. Modifications in chromatin signatures such as H3K4me3 and H3K9ac, which are associated with transcriptionally active genes, can lead to genomic instability and perturb the expression of gene sets associated with oncogenic processes. In order to further elucidate early pre-tumorigenic epigenetic molecular events driving CRC, we integrated diverse, genome-wide, epigenetic inputs (by high throughput sequencing of RNA, H3K4me3, and H3K9ac) and compared differentially expressed transcripts (DE) and enriched regions (DER) in an in-vivo rat colon cancer progression model. Carcinogen (AOM) effects were detected genome-wide at the RNA (116 DE genes), K9ac (49 DERs including 24 genes) and K4me3 (7678 DERs including 3792 genes) level. RNA-seq differential expression and pathway analysis indicated that interferon-associated innate immune responses were impacted by AOM exposure. Despite extensive associations between K4me3 DERs and colon tumorigenesis (1210 genes were linked to colorectal carcinoma) including FOXO3, GNAI2, H2AFX, MSH2, NR3C1, PDCD4 and VEGFA, these changes were not reflected at the RNA gene expression level during early cancer progression. Collectively, our results indicate that carcinogen-induced changes in gene K4me3 DERs are harbingers of future transcriptional events, which drive malignant transformation of the colon.
The galactose-induced decrease in phosphate levels leads to toxicity in yeast models of galactosemia by Caio M. Machado; Evandro A. De-Souza; Ana Luiza F.V. De-Queiroz; Felipe S.A. Pimentel; Guilherme F.S. Silva; Fabio M. Gomes; Mónica Montero-Lomelí; Claudio A. Masuda (1403-1409).
Classic galactosemia is an inborn error of metabolism caused by deleterious mutations in the GALT gene. A number of evidences indicate that the galactose-1-phosphate accumulation observed in patient cells is a cause of toxicity in this disease. Nevertheless, the consequent molecular events caused by the galactose-1-phosphate accumulation remain elusive. Here we show that intracellular inorganic phosphate levels decreased when yeast models of classic galactosemia were exposed to galactose. The decrease in phosphate levels is probably due to the trapping of phosphate in the accumulated galactose-1-phosphate since the deletion of the galactokinase encoding gene GAL1 suppressed this phenotype. Galactose-induced phosphate depletion caused an increase in glycogen content, an expected result since glycogen breakdown by the enzyme glycogen phosphorylase is dependent on inorganic phosphate. Accordingly, an increase in intracellular phosphate levels suppressed the galactose effect on glycogen content and conferred galactose tolerance to yeast models of galactosemia. These results support the hypothesis that the galactose-induced decrease in phosphate levels leads to toxicity in galactosemia and opens new possibilities for the development of better treatments for this disease.
Keywords: Galactosemia; Phosphate trapping; Galactose-1-phosphate; Saccharomyces cerevisiae; Lithium; Unfolded protein response;
Copper therapy reduces intravascular hemolysis and derepresses ferroportin in mice with mosaic mutation (Atp7a mo-ms ): An implication for copper-mediated regulation of the Slc40a1 gene expression by Małgorzata Lenartowicz; Rafał R. Starzyński; Aneta Jończy; Robert Staroń; Justyna Antoniuk; Wojciech Krzeptowski; Paweł Grzmil; Aleksandra Bednarz; Olga Pierzchała; Mateusz Ogórek; Zenon Rajfur; Zbigniew Baster; Paweł Lipiński (1410-1421).
Mosaic mutant mice displaying functional dysfunction of Atp7a copper transporter (the Menkes ATPase) are an established animal model of Menkes disease and constitute a convenient tool for investigating connections between copper and iron metabolisms. This model allows to explore changes in iron metabolism in suckling mutant mice suffering from systemic copper deficiency as well as in young and adult ones undergone copper therapy, which reduces lethal effect of the Atp7a gene mutation. Our recent study demonstrated that 14-day-old mosaic mutant males display blood cell abnormalities associated with intravascular hemolysis, and show disturbances in the functioning of the hepcidin-ferroportin regulatory axis, which controls systemic iron homeostasis. We thus aimed to check whether copper supplementation recovers mutants from hemolytic insult and rebalance systemic iron regulation. Copper supplementation of 14-day-old mosaic mutants resulted in the reestablishment of hematological status, attenuation of hepicidin and concomitant induction of the iron exporter ferroportin/Slc40a1 expression in the liver, down-regulated in untreated mutants. Interestingly, treatment of wild-type males with copper, induced hepcidin-independent up-regulation of ferroportin protein level in hepatic macrophages in both young and adult (6-month-old) animals. Stimulatory effect of copper on ferroportin mRNA and protein levels was confirmed in bone marrow-derived macrophages isolated from both wild-type and mosaic mutant males. Our study indicates that copper is an important player in the regulation of the Slc40a1 gene expression.
Keywords: ATP7A; Copper supplementation; Ferroportin, iron metabolism; Menkes disease; Mottled mice;
Down-regulation of the mitochondrial aspartate-glutamate carrier isoform 1 AGC1 inhibits proliferation and N-acetylaspartate synthesis in Neuro2A cells by Emanuela Profilo; Luis Emiliano Peña-Altamira; Mariangela Corricelli; Alessandra Castegna; Alberto Danese; Gennaro Agrimi; Sabrina Petralla; Giulia Giannuzzi; Vito Porcelli; Luigi Sbano; Carlo Viscomi; Francesca Massenzio; Erika Mariana Palmieri; Carlotta Giorgi; Giuseppe Fiermonte; Marco Virgili; Luigi Palmieri; Massimo Zeviani; Paolo Pinton; Barbara Monti; Ferdinando Palmieri; Francesco Massimo Lasorsa (1422-1435).
The mitochondrial aspartate-glutamate carrier isoform 1 (AGC1) catalyzes a Ca2 +-stimulated export of aspartate to the cytosol in exchange for glutamate, and is a key component of the malate-aspartate shuttle which transfers NADH reducing equivalents from the cytosol to mitochondria. By sustaining the complete glucose oxidation, AGC1 is thought to be important in providing energy for cells, in particular in the CNS and muscle where this protein is mainly expressed. Defects in the AGC1 gene cause AGC1 deficiency, an infantile encephalopathy with delayed myelination and reduced brain N-acetylaspartate (NAA) levels, the precursor of myelin synthesis in the CNS. Here, we show that undifferentiated Neuro2A cells with down-regulated AGC1 display a significant proliferation deficit associated with reduced mitochondrial respiration, and are unable to synthesize NAA properly. In the presence of high glutamine oxidation, cells with reduced AGC1 restore cell proliferation, although oxidative stress increases and NAA synthesis deficit persists. Our data suggest that the cellular energetic deficit due to AGC1 impairment is associated with inappropriate aspartate levels to support neuronal proliferation when glutamine is not used as metabolic substrate, and we propose that delayed myelination in AGC1 deficiency patients could be attributable, at least in part, to neuronal loss combined with lack of NAA synthesis occurring during the nervous system development.
Keywords: Mitochondrial aspartate/glutamate carrier; AGC1 deficiency; Brain hypomyelination; Neurodegenerative disorders; N-Acetylaspartate synthesis;
Knockdown of sestrin2 increases pro-inflammatory reactions and ER stress in the endothelium via an AMPK dependent mechanism by Hwan-Jin Hwang; Tae Woo Jung; Ju-Hee Choi; Hyun Jung Lee; Hye Soo Chung; Ji A Seo; Sin Gon Kim; Nan Hee Kim; Kyung Mook Choi; Dong Seop Choi; Sei Hyun Baik; Hye Jin Yoo (1436-1444).
Sestrin2 (sesn2) has recently gained attention as an important regulator for various metabolic disorders. Sesn2 is involved in AMP-activated protein kinase (AMPK) activation, which leads to anti-inflammatory and anti-oxidative responses. However, the role of sesn2 in the endothelium has not yet been clarified.To evaluate sesn2-mediated anti-atherosclerotic effects, siRNA to silence sesn2 expression was introduced to human umbilical vein endothelial cells (HUVECs), THP-1 cells and C57BL/6 mice. Lipopolysaccharide (LPS) was administrated to sesn2-knockdown cells and mice to induce atherosclerotic signals.Knockdown of sesn2 was involved with atherosclerotic reactions caused by LPS treatment through decrease of AMPK phosphorylation. In sesn2-knockdown HUVECs and THP-1 cells, LPS-mediated nuclear factor kappa B (NF-κB) phosphorylation and secretion of pro-inflammatory cytokines were both significantly increased. In HUVECs, expression of adhesion molecules and LPS-stimulated adhesion of THP-1 cells to the endothelium were significantly increased after sesn2-knockdown. Furthermore, LPS-induced reactive oxygen species (ROS) production, endoplasmic reticulum (ER) stress, and cell toxicity were all significantly elevated after sesn2-knockdown in HUVECs. Interestingly, all these pro-atherosclerotic effects were fully abrogated by treatment with an AMPK activator. In aortic tissue samples from C57BL/6 mice, sesn2-knockdown using siRNA oligomers resulted in reduced AMPK phosphorylation and induction of LPS-mediated NF-κB phosphorylation, leading to up-regulation of adhesion molecules and ER stress-related signaling.Knockdown of sesn2 aggravates atherosclerotic processes by increasing pro-inflammatory reactions and ER stress in the endothelium via an AMPK-dependent mechanism, suggesting that sesn2 might be a novel therapeutic target for atherosclerosis.
Keywords: Sesn2; AMPK; Pro-inflammatory reactions; ER stress; Atherosclerosis; Endothelium;
Integrative network analysis reveals time-dependent molecular events underlying left ventricular remodeling in post-myocardial infarction patients by Florence Pinet; Marie Cuvelliez; Thomas Kelder; Philippe Amouyel; Marijana Radonjic; Christophe Bauters (1445-1453).
To elucidate the time-resolved molecular events underlying the LV remodeling (LVR) process, we developed a large-scale network model that integrates the 24 molecular variables (plasma proteins and non-coding RNAs) collected in the REVE-2 study at four time points (baseline, 1 month, 3 months and 1 year) after MI. The REVE-2 network model was built by extending the set of REVE-2 variables with their mechanistic context based on known molecular interactions (1310 nodes and 8639 edges). Changes in the molecular variables between the group of patients with high LVR (> 20%) and low LVR (< 20%) were used to identify active network modules within the clusters associated with progression of LVR, enabling assessment of time-resolved molecular changes. Although the majority of molecular changes occur at the baseline, two network modules specifically show an increasing number of active molecules throughout the post-MI follow up: one involved in muscle filament sliding, containing the major troponin forms and tropomyosin proteins, and the other associated with extracellular matrix disassembly, including matrix metalloproteinases, tissue inhibitors of metalloproteinases and laminin proteins. For the first time, integrative network analysis of molecular variables collected in REVE-2 patients with known molecular interactions allows insight into time-dependent mechanisms associated with LVR following MI, linking specific processes with LV structure alteration. In addition, the REVE-2 network model provides a shortlist of prioritized putative novel biomarker candidates for detection of LVR after MI event associated with a high risk of heart failure and is a valuable resource for further hypothesis generation.Graphical abstractDisplay Omitted
Keywords: Left ventricule remodeling; echocardiography; biomarkers; system biology;
Nicotine slows down oligomerisation of α-synuclein and ameliorates cytotoxicity in a yeast model of Parkinson's disease by Jay Kardani; Ratnika Sethi; Ipsita Roy (1454-1463).
Several retrospective epidemiological reports have indicated an inverse correlation between smoking and development of Parkinson's disease (PD). This has mostly been attributed to the neuroprotective role of nicotine in stimulating nicotinic acetylcholine receptors and dopaminergic neurons which are damaged in PD. One of the characteristic features of PD is the intraneuronal deposition of globular inclusions of the intrinsically disordered protein α-synuclein as Lewy bodies. Using in vitro and the well-validated yeast cell models, we show that nicotine also exerts a beneficial effect on aggregation of α-synuclein. The alkaloid increases the lag time of the nucleation step and reduces the build-up of the more toxic oligomeric species in a concentration-dependent manner. This results in lower oxidative stress in the cell, reduced cytotoxicity and increased cell survival. Structural studies using CD spectroscopy and fluorescence quenching showed that α-synuclein forms a transient complex with nicotine, distorting its native structure and altering its aggregation landscape such that the formation of oligomers is inhibited. As soluble oligomers are believed to modulate the mechanism of PD pathogenesis mainly by formation of pores in neuronal membranes, resulting in leaching of vital components of the cytoplasm with deleterious effects for the cell, our results provide a mechanistic rationale for the observed beneficial role of nicotine on the progression of the disease.Display Omitted
Keywords: Amyloid fibrils; Fibrillation; Lag time; Nicotine; Oligomerisation;
Deletion of CD73 in mice leads to aortic valve dysfunction by P. Zukowska; B. Kutryb-Zajac; A. Jasztal; M. Toczek; M. Zabielska; T. Borkowski; Z. Khalpey; R.T. Smolenski; E.M. Slominska (1464-1472).
Aortic stenosis is known to involve inflammation and thrombosis. Changes in activity of extracellular enzyme - ecto-5′-nucleotidase (referred also as CD73) can alter inflammatory and thrombotic responses. This study aimed to evaluate the effect of CD73 deletion in mice on development of aortic valve dysfunction and to compare it to the effect of high-fat diet.Four groups of mice (normal-diet Wild Type (WT), high-fat diet WT, normal diet CD73 −/−, high-fat diet CD73 −/−) were maintained for 15 weeks followed by echocardiographic analysis of aortic valve function, measurement of aortic surface activities of nucleotide catabolism enzymes as well as alkaline phosphatase activity, mineral composition and histology of aortic valve leaflets.CD73 −/− knock out led to an increase in peak aortic flow (1.06 ± 0.26 m/s) compared to WT (0.79 ± 0.26 m/s) indicating obstruction. Highest values of peak aortic flow (1.26 ± 0.31 m/s) were observed in high-fat diet CD73 −/− mice. Histological analysis showed morphological changes in CD73 −/− including thickening and accumulation of dark deposits, proved to be melanin. Concentrations of Ca2 +, Mg2 + and PO4 3 − in valve leaflets were elevated in CD73 −/− mice. Alkaline phosphatase (ALP) activity was enhanced after ATP treatment and reduced after adenosine treatment in aortas incubated in osteogenic medium. AMP hydrolysis in CD73 −/− was below 10% of WT. Activity of ecto-adenosine deaminase (eADA), responsible for adenosine deamination, in the CD73 −/− was 40% lower when compared to WT.Deletion of CD73 in mice leads to aortic valve dysfunction similar to that induced by high-fat diet suggesting important role of this surface protein in maintaining heart valve integrity.Display Omitted
Keywords: Adenosine; Ecto-5′-nucleotidase; Aortic stenosis; Inflammation; CD73 knock-out mice;
Experimental Guillain-Barre syndrome induced by immunization with gangliosides: Keyhole limpet hemocyanin is required for disease triggering by Samanta C. Funes; María Eugenia Chiari; Romina Comín; Fernando J. Irazoqui; Gustavo A. Nores (1473-1478).
An experimental model of Guillain-Barré Syndrome has been established in recent years. Rabbits develop disease upon immunization with a single dose of an emulsion containing bovine brain gangliosides, KLH and complete Freund's adjuvant. Within a period of four to ten weeks after immunization, they began to produce anti-ganglioside IgG-antibodies first, and to show clinical signs of neuropathy afterwards. In addition to gangliosides, KLH is a requirement for antibody production and disease triggering. Although KLH is commonly used as an immunological carrier protein, an anti-KLH-specific immune response was necessary for induction of both events. KLH is a glycoprotein carrying most of the immunogenicity in its glycan moiety. Between 20% to 80% of anti-ganglioside IgG-antibodies present in sick rabbit sera cross-reacted with KLH, indicating that both immune responses are related. The terminal Gal-ß(1,3)-GalNAc glycan (present in gangliosides and KLH) is proposed as “key” antigenic determinant involved in inducing the anti-ganglioside immune response. These results are discussed in the context of the “binding site drift” hypothesis.Display Omitted
Keywords: Anti-ganglioside antibodies; KLH; Guillain-Barré syndrome; Experimental model; Binding site drift;
Down-regulation of islet amyloid polypeptide expression induces death of human annulus fibrosus cells via mitochondrial and death receptor pathways by Xinghuo Wu; Kun Wang; Wenbin Hua; Shuai Li; Xianzhe Liu; Wei Liu; Yu Song; Yukun Zhang; Zengwu Shao; Cao Yang (1479-1491).
Islet amyloid polypeptide (IAPP) exerts its biological effects by participating in the regulation of glucose metabolism and cell apoptosis. The main goal of the present study was to investigate the expression of IAPP in degenerated intervertebral disc tissue and IAPP's modulation of extracellular matrix (ECM) catabolic and anabolic genes in human AF cells. We found that the expression of IAPP, the calcitonin receptor, and receptor activity modifying protein decreased considerably in AF cells during the progression of intervertebral disc degeneration (IDD). Meanwhile, transfection with pLV-siIAPP decreased the expression of IAPP and its receptors and reduced glucose uptake and the expression of aggrecan, Col2A1, and BG. Down-regulation of IAPP also induced a significant increase in reactive oxygen species generation in AF cells, along with a decrease in matrix metalloproteinases and an increase in the concentration of cellular Ca2 + , ultimately leading to death. Further analysis revealed that siIAPP intervention promoted the release of cytochrome c from mitochondria, resulting in the activation of Caspase-3 and Caspase-9. In contrast, significantly decreased expression of Caspase-3 and Caspase-9 was observed in AF cells transfected with pLV-IAPP. The concentrations of Fas and FasL proteins were significantly decreased in AF cells transfected with PLV-IAPP, while activation of the Fas/FasL system and cell death were induced by siIAPP intervention. Mechanistically, AMPK/Akt-mTOR signaling pathways were involved. In conclusion, down-regulation of IAPP expression induces the death of human AF cells via mitochondrial and death receptor pathways, potentially offering a novel therapeutic target for the treatment of IDD.
Keywords: Islet amyloid polypeptide; Apoptosis; Disc degeneration; Extracellular matrix metabolism; Annulus fibrosus;
MDH2 is an RNA binding protein involved in downregulation of sodium channel Scn1a expression under seizure condition by Yong-Hong Chen; Shu-Jing Liu; Mei-Mei Gao; Tao Zeng; Guo-Wang Lin; Na-Na Tan; Hui-Ling Tang; Ping Lu; Tao Su; Wei-Wen Sun; Long-Chang Xie; Yong-Hong Yi; Yue-Sheng Long (1492-1499).
Voltage-gated sodium channel α-subunit type I (NaV1.1, encoded by SCN1A gene) plays a critical role in the excitability of brain. Downregulation of SCN1A expression is associated with epilepsy, a common neurological disorder characterized by recurrent seizures. Here we reveal a novel role of malate dehydrogenase 2 (MDH2) in the posttranscriptional regulation of SCN1A expression under seizure condition. We identified that MDH2 was an RNA binding protein that could bind two of the four conserved regions in the 3′ UTRs of SCN1A. We further showed that knockdown of MDH2 or inactivation of MDH2 activity in HEK-293 cells increased the reporter gene expression through the 3′ UTR of SCN1A, and MDH2 overexpression decreased gene expression by affecting mRNA stability. In the hippocampus of seizure mice, the upregulation of MDH2 expression contributed to the decrease of the NaV1.1 levels at posttranscriptional level. In addition, we showed that the H2O2 levels increased in the hippocampus of the seizure mice, and H2O2 could promote the binding of MDH2 to the binding sites of Scn1a gene, whereas β-mercaptoethanol decreased the binding capability, indicating an important effect of the seizure-induced oxidation on the MDH2-mediated downregulation of Scn1a expression. Taken together, these data suggest that MDH2, functioning as an RNA-binding protein, is involved in the posttranscriptional downregulation of SCN1A expression under seizure condition.
Keywords: MDH2; Scn1a; Seizure; Posttranscriptional regulation; RNA binding protein;
Conformational and thermal characterization of left ventricle remodeling post-myocardial infarction by V. Samouillan; E. Revuelta-López; C. Soler-Botija; J. Dandurand; Aleyda Benitez-Amaro; L. Nasarre; D. de Gonzalo-Calvo; A. Bayes-Genis; C. Lacabanne; V. Llorente-Cortés (1500-1509).
Adverse cardiac remodeling after myocardial infarction (MI) causes impaired ventricular function and heart failure. Histopathological characterization is commonly used to detect the location, size and shape of MI sites. However, the information about chemical composition, physical structure and molecular mobility of peri- and infarct zones post-MI is rather limited. The main objective of this work was to explore the spatiotemporal biochemical and biophysical alterations of key cardiac components post-MI. The FTIR spectra of healthy and remote myocardial tissue shows amides A, I, II and III associated with proteins in freeze-died tissue as major absorptions bands. In infarcted myocardium, the spectrum of these main absorptions was deeply altered. FITR evidenced an increase of the amide A band and the distinct feature of the collagen specific absorption band at 1338 cm− 1 in the infarct area at 21 days post-MI. At 21 days post-MI, it also appears an important shift of amide I from 1646 cm− 1 to 1637 cm− 1 that suggests the predominance of the triple helical conformation in the proteins. The new spectra bands also indicate an increase in proteoglycans, residues of carbohydrates in proteins and polysaccharides in ischemic areas. Thermal analysis indicates a deep increase of unfreezable water/freezable water in peri- and infarcted tissues. In infarcted tissue is evidenced the impairment of myofibrillar proteins thermal profile and the emergence of a new structure. In conclusion, our results indicate a profound evolution of protein secondary structures in association with collagen deposition and reorganization of water involved in the scar maturation of peri- and infarct zones post-MI.
Keywords: Myocardial infarction; FITR; DSC; Physical characterization;
Interleukin-17A exacerbates high-fat diet-induced hepatic steatosis by inhibiting fatty acid β-oxidation by Tianran Shen; Xu Chen; Yanping Li; Xilan Tang; Xinwei Jiang; Chao Yu; Yuanzhu Zheng; Honghui Guo; Wenhua Ling (1510-1518).
There is a growing body of evidence that the interleukin-17A (IL-17A) signaling pathway contributes to the pathogenesis of nonalcoholic fatty liver disease (NAFLD). However, the mechanism by which IL-17A signaling induces hepatocyte injury is unclear. The aim of the present study was to investigate the significance of the IL-17A axis in NAFLD and to explore the role of IL-17A in high-fat diet (HFD)-induced NAFLD in C57BL/6 mice and oleic acid (OA)-induced lipid accumulation in hepatocytes. Firstly, Consistent upregulation of IL-17A was observed in the HFD-induced steatosis mice but not the normal chow-fed control mice. Administration of IL-17A impaired liver function, aggravated hepatic lipid accumulation by inhibiting fatty acid oxidation in the HFD mice. Conversely, inhibition of IL-17A using an anti-IL-17A monoclonal antibody (mAb) significantly attenuated HFD-induced liver injury. Furthermore, IL-17A accelerated hepatic steatosis through activation of the JNK-PPARα pathway in the HFD mice and OA-preloaded hepatocytes.The present study demonstrated that a high fat diet induces IL-17A expression, which exacerbates the progression of NAFLD by inhibiting fatty acid β-oxidation and promoting the accumulation of triglycerides (TG).Display Omitted
Keywords: Fatty acid metabolism; Inflammation; JNK-PPARα; NAFLD;
Novel therapeutic approaches for disease-modification of epileptogenesis for curing epilepsy by Bryan L. Clossen; Doodipala Samba Reddy (1519-1538).
This article describes the recent advances in epileptogenesis and novel therapeutic approaches for the prevention of epilepsy, with a special emphasis on the pharmacological basis of disease-modification of epileptogenesis for curing epilepsy. Here we assess animal studies and human clinical trials of epilepsy spanning 1982–2016. Epilepsy arises from a number of neuronal factors that trigger epileptogenesis, which is the process by which a brain shifts from a normal physiologic state to an epileptic condition. The events precipitating these changes can be of diverse origin, including traumatic brain injury, cerebrovascular damage, infections, chemical neurotoxicity, and emergency seizure conditions such as status epilepticus. Expectedly, the molecular and system mechanisms responsible for epileptogenesis are not well defined or understood. To date, there is no approved therapy for the prevention of epilepsy. Epigenetic dysregulation, neuroinflammation, and neurodegeneration appear to trigger epileptogenesis. Targeted drugs are being identified that can truly prevent the development of epilepsy in at-risk people. The promising agents include rapamycin, COX-2 inhibitors, TRK inhibitors, epigenetic modulators, JAK-STAT inhibitors, and neurosteroids. Recent evidence suggests that neurosteroids may play a role in modulating epileptogenesis. A number of promising drugs are under investigation for the prevention or modification of epileptogenesis to halt the development of epilepsy. Some drugs in development appear rational for preventing epilepsy because they target the initial trigger or related signaling pathways as the brain becomes progressively more prone to seizures. Additional research into the target validity and clinical investigation is essential to make new frontiers in curing epilepsy.
Keywords: Epilepsy; Epileptogenesis; Epigenetics; Neurosteroid; JAK-Stat; Rapamycin;
Mitochondrial DNA maintenance defects by Ayman W. El-Hattab; William J. Craigen; Fernando Scaglia (1539-1555).
The maintenance of mitochondrial DNA (mtDNA) depends on a number of nuclear gene-encoded proteins including a battery of enzymes forming the replisome needed to synthesize mtDNA. These enzymes need to be in balanced quantities to function properly that is in part achieved by exchanging intramitochondrial contents through mitochondrial fusion. In addition, mtDNA synthesis requires a balanced supply of nucleotides that is achieved by nucleotide recycling inside the mitochondria and import from the cytosol. Mitochondrial DNA maintenance defects (MDMDs) are a group of diseases caused by pathogenic variants in the nuclear genes involved in mtDNA maintenance resulting in impaired mtDNA synthesis leading to quantitative (mtDNA depletion) and qualitative (multiple mtDNA deletions) defects in mtDNA. Defective mtDNA leads to organ dysfunction due to insufficient mtDNA-encoded protein synthesis, resulting in an inadequate energy production to meet the needs of affected organs. MDMDs are inherited as autosomal recessive or dominant traits, and are associated with a broad phenotypic spectrum ranging from mild adult-onset ophthalmoplegia to severe infantile fatal hepatic failure. To date, pathogenic variants in 20 nuclear genes known to be crucial for mtDNA maintenance have been linked to MDMDs, including genes encoding enzymes of mtDNA replication machinery (POLG, POLG2, TWNK, TFAM, RNASEH1, MGME1, and DNA2), genes encoding proteins that function in maintaining a balanced mitochondrial nucleotide pool (TK2, DGUOK, SUCLG1, SUCLA2, ABAT, RRM2B, TYMP, SLC25A4, AGK, and MPV17), and genes encoding proteins involved in mitochondrial fusion (OPA1, MFN2, and FBXL4).
Keywords: mitochondrial diseases; mitochondrial DNA (mtDNA); mtDNA depletion syndromes; multiple mtDNA deletions; mitochondrial fusion; mtDNA replication;
A novel mechanism of diabetic vascular endothelial dysfunction: Hypoadiponectinemia-induced NLRP3 inflammasome activation by Jinglong Zhang; Linying Xia; Fen Zhang; Di Zhu; Chao Xin; Helin Wang; Fuyang Zhang; Xian Guo; Yan Lee; Ling Zhang; Shan Wang; Xiong Guo; Chong Huang; Feng Gao; Yi Liu; Ling Tao (1556-1567).
It has been well documented that hypoadiponectinemia is associated with impaired endothelium-dependent vasodilation. However, the exact molecular mechanism which mediates this process has not been fully described. The current study aimed to investigate the role of hypoadiponectinemia-induced NLRP3 inflammasome activation in diabetic vascular endothelial dysfunction and its molecular mechanism. Male adult adiponectin knockout mice and wild type mice were fed with a high fat diet to establish a type 2 diabetic mellitus model. In addition, human umbilical vein endothelial cells (HUVECs) were cultured and subjected to high glucose/high fat (HG/HF). The NLRP3 inflammasome activation was increased in type 2 diabetic mice and treatment of diabetic aortic segments with MCC950, a potent selective inhibitor of NLRP3 inflammasome ex vivo improved endothelial-dependent vasorelaxation. NLRP3 inflammasome activation and vascular endothelial injury were significantly increased in APN-KO mice compared with WT mice in diabetes and MCC950 decreased diabetic vascular endothelial dysfunction to comparable levels in APN-KO mice and WT mice. Adiponectin could decrease NLRP3 inflammasome activation and attenuate endothelial cell injury, which was abolished by NLRP3 inflammasome overexpression. Inhibition of peroxynitrite formation preferentially attenuated NLRP3 inflammasome activation in APN-KO diabetic mice. The current study demonstrated for the first time that hypoadiponectinemia-induced NLRP3 inflammasome activation was a novel mechanism of diabetic vascular endothelial dysfunction.Display Omitted
Keywords: Adiponectin; Endothelial cells; NLRP3 inflammasome; Oxidative/nitrative stress; Type 2 diabetic mellitus;
Sodium restriction modulates innate immunity and prevents cardiac remodeling in a rat model of metabolic syndrome by Bernard Jover; Christelle Reynes; Caroline Rugale; Cyril Reboul; Laura Jeanson; Michel Tournier; Anne Dominique Lajoix; Caroline Desmetz (1568-1574).
In the view of the relationships between excessive sodium intake, immunity and target organ damage, we hypothesized that reduction in dietary sodium would be beneficial in the prevention of cardiac alterations through a restrained local immunity response in a rat model of metabolic syndrome. Sprague-Dawley rats were fed a 60% fructose diet with either a normal sodium (0.64% NaCl) or a low sodium content (< 0.01% NaCl) for 8 weeks. After 4 weeks, rats were infused or not with angiotensin II (200 ng·kg− 1·min− 1, sc) for 4 weeks. Tail-cuff blood pressure was determined in conscious rats. Heart and left ventricle weight, cardiomyocyte size, and cardiac fibrosis were evaluated. We performed a transcriptomic analysis in order to identify differentially regulated cardiac mRNAs between normal and low sodium diets. We validated those results using qPCR and immunohistochemistry.Angiotensin II-induced blood pressure rise was blunted (~ 50%) in the low-sodium fed rats while cardiac hypertrophy and fibrosis were prevented. Transcriptomic analysis revealed 66 differentially regulated genes including 13 downregulated genes under the low sodium diet and implicated in the innate immune response. This was confirmed by reduced cardiac macrophages infiltration under the low sodium diet.Dietary sodium restriction prevents structural alterations of the heart of rats with fructose-induced insulin resistance and angiotensin II-hypertension. The reduction of cardiac inflammation and macrophage infiltration suggests that innate immunity has an important role in the beneficial effect of sodium restriction on cardiac remodeling.
Keywords: Low sodium diet; Cardiac hypertrophy; Fibrosis; Innate immunity; Macrophage; Inflammation;
EGCG ameliorates diet-induced metabolic syndrome associating with the circadian clock by Yashi Mi; Guoyuan Qi; Rong Fan; Xiaohua Ji; Zhigang Liu; Xuebo Liu (1575-1589).
In response to the daily light-dark (LD) cycle, organisms on Earth have evolved with the approximately 24-h endogenous oscillations to coordinate behavioral and physiological processes, including feeding, sleep, and metabolism homeostasis. Circadian desynchrony triggered by an energy-dense diet rich in fats and fructose is intimately connected with a series of metabolic disorders. Previous studies revealed that (−)-Epigallocatechin-3-gallate (EGCG) could mitigate metabolic misalignment; however, only a few reports have focused on its potential effect on directly manipulating circadian rhythms to ameliorate metabolic syndrome. Our goal was to investigate the regulating effect of EGCG treatment on metabolic misalignment triggered by a high-fat and high-fructose diet (HFFD) associating with the circadian clock. Our results indicated that HFFD treatment partially exhibited poor circadian oscillations of the core clock gene and the clock-controlled gene in the liver and fat relative to the control group. EGCG administration may ameliorate the diet-dependent decline in circadian function by controlling the Sirt1-PGC1αloop, implying the existence of an EGCG-entrainable oscillator. Subsequently, reducing fatty acid synthesis and elevating β-oxidation in the liver coupled with the increasing brown adipose tissue (BAT) energy expenditure observed in the EGCG group of mice prevented the adipocyte hypertrophy and fat accumulations common to BAT and white adipose tissue (WAT) derived from the HFFD mice. This study is the first to provide compelling evidences that EGCG may ameliorate diet-induced metabolic misalignment by regulating the rhythmic expression of the circadian clock genes in the liver and fat.Display Omitted
Keywords: (−)-Epigallocatechin-3-gallate (EGCG); Circadian rhythm; Metabolic syndrome; Insulin resistance;
Metabolomic profiling of lung function in Costa-Rican children with asthma by Rachel S. Kelly; Yamini Virkud; Rachel Giorgio; Juan C. Celedón; Scott T. Weiss; Jessica Lasky-Su (1590-1595).
The development of novel therapeutics and treatment regimens for the management of asthma is hindered by an incomplete understanding of its heterogeneous nature and pathophysiology. Metabolomics can provide an integrated and global profile of a biological system in a dysregulated state, making it a valuable tool to identify biomarkers along the disease development pathway and to understand the biological mechanisms driving that pathway.Liquid chromatography-mass spectrometry metabolomic profiling was conducted on plasma samples provided at recruitment for 380 children with asthma from the ‘Genetic Epidemiology of Asthma in Costa Rica Cohort’. Metabolites associated with three clinical characteristics of asthma severity (i) airway hyper-responsiveness (AHR) (ii) percent-predicted forced expiratory volume in one second/forced vital capacity ratio (FEV1/FVC), and (iii) FEV1/FVC post-bronchodilator were identified and their discriminatory ability assessed. Metabolite set enrichment analyses was applied to explore the biology underlying these relationships.AHR was associated (p < 0.05) with 91 of 574 metabolites (15.9%), FEV1/FVC pre-bronchodilator with 102(17.8%), and FEV1/FVC post-bronchodilator with 155 (27.0%). The findings suggest that these characteristics capture some common and some distinct phenotypic aspects of lung function; glycerophospholipid, linoleic acid and pyrimidine metabolism were common to all three characteristics. The corresponding metabolomic profiles showed moderate but robust discriminatory ability.The results confirm the existence of an asthma severity metabolome. However, differences in the metabolomic profiles of the three lung function characteristics studied, suggest that refinement of both phenotype classification and metabolite selection should be a priority as the field of asthma metabolomics progresses.
Keywords: Metabolomics; Asthma; Lung function; FEV1/FVC ratio; Airway hyperresponsiveness;
Reduced bioavailable manganese causes striatal urea cycle pathology in Huntington's disease mouse model by Terry Jo V. Bichell; Michal Wegrzynowicz; K. Grace Tipps; Emma M. Bradley; Michael A. Uhouse; Miles Bryan; Kyle Horning; Nicole Fisher; Karrie Dudek; Timothy Halbesma; Preethi Umashanker; Andrew D. Stubbs; Hunter K. Holt; Gunnar F. Kwakye; Andrew M. Tidball; Roger J. Colbran; Michael Aschner; M. Diana Neely; Alba Di Pardo; Vittorio Maglione; Alexander Osmand; Aaron B. Bowman (1596-1604).
Huntington's disease (HD) is caused by a mutation in the huntingtin gene (HTT), resulting in profound striatal neurodegeneration through an unknown mechanism. Perturbations in the urea cycle have been reported in HD models and in HD patient blood and brain. In neurons, arginase is a central urea cycle enzyme, and the metal manganese (Mn) is an essential cofactor. Deficient biological responses to Mn, and reduced Mn accumulation have been observed in HD striatal mouse and cell models. Here we report in vivo and ex vivo evidence of a urea cycle metabolic phenotype in a prodromal HD mouse model. Further, either in vivo or in vitro Mn supplementation reverses the urea-cycle pathology by restoring arginase activity. We show that Arginase 2 (ARG2) is the arginase enzyme present in these mouse brain models, with ARG2 protein levels directly increased by Mn exposure. ARG2 protein is not reduced in the prodromal stage, though enzyme activity is reduced, indicating that altered Mn bioavailability as a cofactor leads to the deficient enzymatic activity. These data support a hypothesis that mutant HTT leads to a selective deficiency of neuronal Mn at an early disease stage, contributing to HD striatal urea-cycle pathophysiology through an effect on arginase activity.Display Omitted
Keywords: Huntington's; Neurodegeneration; Manganese; Arginase; Striatum; Urea;
Modification of the fatty acid composition of an obesogenic diet improves the maternal and placental metabolic environment in obese pregnant mice by Martina Gimpfl; Jan Rozman; Maik Dahlhoff; Raphaela Kübeck; Andreas Blutke; Birgit Rathkolb; Martin Klingenspor; Martin Hrabě de Angelis; Soner Öner-Sieben; Annette Seibt; Adelbert A. Roscher; Eckhard Wolf; Regina Ensenauer (1605-1614).
Peri-conceptional exposure to maternal obesogenic nutrition is associated with in utero programming of later-life overweight and metabolic disease in the offspring. We aimed to investigate whether dietary intervention with a modified fatty acid quality in an obesogenic high-calorie (HC) diet during the preconception and gestational phases can improve unfavourable effects of an adipogenic maternal environment. In NMRI mice, peri-conceptional and gestational obesity was induced by feeding a HC diet (controls), and they were compared with dams on a fat-modified (Fat-mod) HC diet of the same energy content but enriched with medium-chain fatty acids (MCFAs) and adjusted to a decreased ratio of n-6 to n-3 long-chain polyunsaturated fatty acids (LC-PUFAs). Effects on maternal and placental outcomes at delivery (day 17.5 post coitum) were investigated. Despite comparable energy assimilation between the two groups of dams, the altered fatty acid composition of the Fat-mod HC diet induced lower maternal body weight, weights of fat depots, adipocyte size, and hepatic fat accumulation compared to the unmodified HC diet group. Further, there was a trend towards lower fasting glucose, insulin and leptin concentrations in dams fed the Fat-mod HC diet. Phenotypic changes were accompanied by inhibition of transcript and protein expression of genes involved in hepatic de novo lipogenesis comprising PPARG2 and its target genes Fasn, Acaca, and Fabp4, whereas regulation of other lipogenic factors (Srebf1, Nr1h3, Abca1) appeared to be more complex. The modified diet led to a sex-specific placental response by upregulating PPARG-dependent fatty acid transport gene expression in female versus male placentae. Qualitative modification of the fatty acid spectrum of a high-energy maternal diet, using a combination of both MCFAs and n-3 LC-PUFAs, seems to be a promising interventional approach to ameliorate the adipogenic milieu of mice before and during gestation.
Keywords: Pregnancy; Obesity; Nutrition; Medium-chain fatty acids; n-3 fatty acids; Placenta;
RNF8 identified as a co-activator of estrogen receptor α promotes cell growth in breast cancer by Shengli Wang; Hao Luo; Chunyu Wang; Hongmiao Sun; Ge Sun; Ning Sun; Kai Zeng; Huijuan Song; Renlong Zou; Tingting Zhou; Rijiao Cong; Wei Liu; Lei Yang; Da Li; Xin Zhou; Xinping Zhong; Lin Lin; Jiao Jiao; Guangqi Yan; Xue Wang; Xiaojie Min; Liu Cao; Yue Zhao (1615-1628).
The ring finger protein 8 (RNF8), a key component of protein complex crucial for DNA-damage response, consists of a forkhead-associated (FHA) domain and a really interesting new gene (RING) domain that enables it to function as an E3 ubiquitin ligase. However, the biological functions of RNF8 in estrogen receptor α (ERα)-positive breast cancer and underlying mechanisms have not been fully defined. Here, we have explored RNF8 as an associated partner of ERα in breast cancer cells, and co-activates ERα-mediated transactivation. Accordingly, RNF8 depletion inhibits the expression of endogenous ERα target genes. Interestingly, our results have demonstrated that RNF8 increases ERα stability at least partially if not all via triggering ERα monoubiquitination. RNF8 functionally promotes breast cancer cell proliferation. RNF8 is highly expressed in clinical breast cancer samples and the expression of RNF8 positively correlates with that of ERα. Up-regulation of ERα-induced transactivation by RNF8 might contribute to the promotion of breast cancer progression by allowing enhancement of ERα target gene expression. Our study describes RNF8 as a co-activator of ERα increases ERα stability via post-transcriptional pathway, and provides a new insight into mechanisms for RNF8 to promote cell growth of ERα-positive breast cancer.
Keywords: RNF8; Co-activator; Estrogen receptor α; Ubiquitination; Breast cancer;
Heat shock protein 22 (HSPB8) reduces the migration of hepatocellular carcinoma cells through the suppression of the phosphoinositide 3-kinase (PI3K)/AKT pathway by Rie Matsushima-Nishiwaki; Hidenori Toyoda; Reika Takamatsu; Eisuke Yasuda; Seiji Okuda; Atsuyuki Maeda; Yuji Kaneoka; Naoki Yoshimi; Takashi Kumada; Osamu Kozawa (1629-1639).
Small heat shock proteins (HSPs) regulate a variety of cell functions. Among them, HSP22 and HSP20 are recognized to be ubiquitously expressed in various tissues. With regard to hepatocellular carcinoma (HCC) cells, we previously reported that phosphorylated HSP20 plays a suppressive role in transforming growth factor (TGF)-α-induced cell migration and invasion. In the present study, we investigated whether or not HSP22 is implicated in HCC cell migration. We detected HSP22 protein expression both in human HCC tumor (189.9 ± 68.4 ng/mg protein) and the adjacent non-tumor liver tissues (167.9 ± 94.6 ng/mg protein). The cases of low-quantity HSP22 protein level group (88.3 ≧ ng/mg protein, the optimum cut-off value of HSP22) were increased in tumor tissues compared with the adjacent non-tumor tissues. The migration of human HCC-derived HuH-7 cells stimulated by TGF-α or hepatocyte growth factor (HGF) was significantly enhanced by the knockdown of HSP22 expression. Down-regulation of HSP22 protein in the cells markedly strengthened the AKT phosphorylation induced by TGF-α or HGF. Inhibitors of the phosphoinositide 3-kinase (PI3K)/AKT pathway, which suppressed the TGF-α-induced migration, significantly reduced the amplification by HSP22 knockdown. PI3K but not AKT was coimmunoprecipitated with HSP22 in HuH-7 cells. In addition, in human HCC tissues, a significantly lower HSP22 protein level in tumor tissues than in adjacent non-tumor tissues was observed more frequently in cases of moderately or poorly differentiated HCC than well-differentiated HCC. Taken together, our results strongly suggest that HSP22 represses HCC progression, especially HCC cell migration, by the down-regulation of the PI3K/AKT signaling pathway.
Keywords: Heat shock protein 22; Hepatocellular carcinoma; Phosphoinositide 3-kinase; AKT; Migration; Growth factor;
Identification of a molecular signaling gene-gene regulatory network between GWAS susceptibility genes ADTRP and MIA3/TANGO1 for coronary artery disease by Chunyan Luo; Fan Wang; Xiang Ren; Tie Ke; Chengqi Xu; Bo Tang; Subo Qin; Yufeng Yao; Qiuyun Chen; Qing Kenneth Wang (1640-1653).
Coronary artery disease (CAD) is the leading cause of death worldwide. GWAS have identified > 50 genomic loci for CAD, including ADTRP and MIA3/TANGO1. However, it is important to determine whether the GWAS genes form a molecular network. In this study, we have uncovered a novel molecular network between ADTRP and MIA3/TANGO1 for the pathogenesis of CAD. We showed that knockdown of ADTRP expression markedly down-regulated expression of MIA3/TANGO1. Mechanistically, ADTRP positively regulates expression of PIK3R3 encoding the regulatory subunit 3 of PI3K, which leads to activation of AKT, resulting in up-regulation of MIA3/TANGO1. Both ADTRP and MIA3/TANGO1 are involved in endothelial cell (EC) functions relevant to atherosclerosis. Knockdown of ADTRP expression by siRNA promoted oxidized-LDL-mediated monocyte adhesion to ECs and transendothelial migration of monocytes, inhibited EC proliferation and migration, and increased apoptosis, which was reversed by expression of constitutively active AKT1 and MIA3/TANGO1 overexpression, while the over-expression of ADTRP in ECs blunted these processes. Knockdown of MIA3/TANGO1 expression also promoted monocyte adhesion to ECs and transendothelial migration of monocytes, and vice versa for overexpression of MIA3/TANGO1. We found that ADTRP negatively regulates the levels of collagen VII and ApoB in HepG2 and endothelial cells, which are downstream regulatory targets of MIA3/TANGOI. In conclusion, we have uncovered a novel molecular signaling pathway for the pathogenesis of CAD, which involves a novel gene-gene regulatory network. We show that ADTRP positively regulates PIK3R3 expression, which leads to activation of AKT and up-regulation of MIA3/TANGO1, thereby regulating endothelial cell functions directly relevant to atherosclerosis.
Keywords: Coronary artery disease (CAD); Myocardial infarction (MI); ADTRP; MIA3 (TANGO1); AKT; PI3K; Collagen VII; ApoB;
Elatoside C protects against ox-LDL-induced HUVECs injury by FoxO1-mediated autophagy induction by Yun Luo; Xiangbao Meng; Ping Zhou; Shan Lu; Meng Qin; Xudong Xu; Guibo Sun; Xiaobo Sun (1654-1665).
The vascular endothelial cell injury induced by oxidized low-density lipoprotein (ox-LDL) is a major contributing factor to the pathogenesis of atherosclerosis. Elatoside C (EsC), a natural saponin isolated from Longya Aralia chinensis L., possesses anti-oxidative activity; however, there is still no report indicating that EsC protects against ox-LDL-induced endothelial cell injury and the exact mechanisms of this protection. Recently, autophagy has attracted extensive attention on the basis of its ability to modulate cell survival. Thus, we determined the role of autophagy in the protective effects of EsC against ox-LDL-induced human umbilical vein endothelial cells (HUVECs). Our results demonstrated that EsC pretreatment reduced ox-LDL-induced HUVECs oxidative injury, increased the number of autophagosomes and modulated the expression of autophagy related proteins. Moreover, autophagy inhibitor 3-methyladenine, chloroquine and BECN1 siRNA obviously abolished the anti-oxidative effects of EsC. Furthermore, our data indicated that EsC significantly increased nuclear FoxO1 expression level and FoxO1 siRNA markedly attenuated the protective effects of EsC. In conclusion, EsC attenuated ox-LDL-induced HUVECs injury by inducing autophagy via increasing FoxO1 expression level. EsC is thus considered as a potential drug for the treatment of atherosclerosis.
Keywords: Autophagy; Apoptosis; Elatoside C; Ox-LDL; FoxO1; HUVECs;
Spastin regulates VAMP7-containing vesicles trafficking in cortical neurons by C. Plaud; V. Joshi; M. Marinello; D. Pastré; T. Galli; P.A. Curmi; A. Burgo (1666-1677).
Alteration of axonal transport has emerged as a common precipitating factor in several neurodegenerative disorders including Human Spastic Paraplegia (HSP). Mutations of the SPAST (SPG4) gene coding for the spastin protein account for 40% of all autosomal dominant uncomplicated HSP. By cleaving microtubules, spastin regulates several cellular processes depending on microtubule dynamics including intracellular membrane trafficking. Axonal transport is fundamental for the viability of motor neurons which often have very long axons and thus require efficient communication between the cell body and its periphery. Here we found that the anterograde velocity of VAMP7 vesicles, but not that of VAMP2, two vesicular-SNARE proteins implicated in neuronal development, is enhanced in SPG4-KO neurons. We showed that this effect is associated with a slight increase of the level of acetylated tubulin in SPG4-KO neurons and correlates with an enhanced activity of kinesin-1 motors. Interestingly, we demonstrated that an artificial increase of acetylated tubulin by drugs reproduces the effect of Spastin KO on VAMP7 axonal dynamics but also increased its retrograde velocity. Finally, we investigated the effect of microtubule targeting agents which rescue axonal swellings, on VAMP7 and microtubule dynamics. Our results suggest that microtubule stabilizing agents, such as taxol, may prevent the morphological defects observed in SPG4-KO neurons not simply by restoring the altered anterograde transport to basal levels but rather by increasing the retrograde velocity of axonal cargoes.
Keywords: Human Spastic Paraplegia; SPG4; Axonal transport; VAMP7; Microtubule targeting agents;
MicroRNA-130b transcriptionally regulated by histone H3 deacetylation renders Akt ubiquitination and apoptosis resistance to 6-OHDA by Liang Xu; Yu Jia; Xiang-Hong Yang; Fang Han; Yang Zheng; Yin Ni; Xu Chen; Jun Hong; Jing-Quan Liu; Qian Li; Ren-Hua Sun; Shi-Jing Mo (1678-1689).
Apoptosis of DA neurons is a contributing cause of disability and death for Parkinson’s disease (PD). Akt may become a potential therapeutic target for PD since Akt has been deactivated during DA neuron apoptosis. We previously demonstrated that Akt confers apoptosis resistance against 6-OHDA in DA neuron-like PC12 cells, yet the underlying mechanisms accounted for this are not fully understood. Here we report that microRNA-130b (miR-130b)-dependent and cylindromatosis (CYLD) repression-mediated Akt ubiquitination renders apoptosis resistance of PC12 cells to 6-OHDA, which elicits histone H3 deacetylation-induced transcriptional downregulation of miR-130b vice versa. CYLD deficiency ubiquitinates Akt at Lys63, thereby phosphorylating Akt and antagonizing 6-OHDA-initiated apoptosis. MiR-130b targetedly represses CYLD and increases apoptosis resistance to 6-OHDA. CYLD repression by miR-130b restores Akt ubiquitination and activation, GSK3β and FoxO3a phosphorylation, FoxO3a removal from Bim promoter as well as Bim downregulation during 6-OHDA administration. CYLD deficiency-mediated Akt activation is instrumental for the apoptosis-resistant phenotypes of miR-130b. In addition, 6-OHDA transcriptionally downregulates miR-130b through recruitment of HDAC3 at the promoter. Furthermore, EPO potentiates the ability of miR-130b to activate Akt and augment apoptosis resistance. Our findings identify the apoptosis-resistant function of miR-130b and suggest that histone H3 deacetylation plays a pivotal role in regulating miR-130b transcription in response to 6-OHDA.
Keywords: MicroRNA; Akt ubiquitination; Histone H3 deacetylation; Apoptosis;
A common polymorphism decreases LRP1 mRNA stability and is associated with increased plasma factor VIII levels by Jiann-Der Lee; Kuang-Ming Hsiao; Pey-Jium Chang; Chih-Cheng Chen; Ya-Wen Kuo; Yen-Chu Huang; Huan-Lin Hsu; Ya-Hui Lin; Chih-Ying Wu; Ying-Chih Huang; Meng Lee; Chia-Yu Hsu; Yi-Ting Pan; Chih-Yu Kuo; Chun-Hsien Lin (1690-1698).
The low-density lipoprotein receptor-related protein 1 (LRP1) gene is associated with increased levels of plasma factor VIII (FVIII). We aimed to explore eight functional genetic LRP1 variants for their potential roles in regulating FVIII levels and acute ischemic stroke (AIS). This genetic association study enrolled 192 patients with AIS and 134 controls. There were no significant differences in the genetic frequency of the eight functional single-nucleotide polymorphisms (SNPs) between the control and AIS groups. However, while analyzing the association between the eight SNPs and plasma FVIII levels, subjects with T/T genotype of rs1800137 (vs. CC + CT) were found to be associated with higher FVIII levels (23.5 IU/dL; 95% confidence interval, 7.4–39.5 IU/dL; P = 0.0044) after adjusting for age, gender, estimated glomerular filtration rate, O blood type, inflammatory state, and body mass index.An analysis of the mRNA stability and abundance was designed and performed using minigene system transfected into HepG2 cells to assess the possible differences in mRNA stabilities between rs1800137 CC (rs1800137C) and TT (rs1800137T) genotypes. Site-directed mutagenesis revealed that rs1800137T accounts for the observed decrease in mRNA stability. The SNP rs1800137, located in exon 8, has been identified as an exon-splicing enhancer in silico. However, alternative splicing of LRP1 without inclusion of exon 8 was not identified. In transfected HepG2 cells, cycloheximide slowed down the degradation of the rs1800137T-containing minigene. These results demonstrate that synonymous SNP rs1800137 can lead to increased plasma FVIII levels due to decreased mRNA stability via translation-dependent mRNA degradation associated with codon optimality.Display Omitted
Keywords: Low density lipoprotein receptor-related protein 1; Codon optimality; Genetic polymorphism; Factor VIII; Messenger RNA stability;