Literature-based discovery of diabetes- and ROS-related targets

Abstract Background Reactive oxygen species (ROS) are known mediators of cellular damage in multiple diseases including diabetic complications. Despite its importance, no comprehensive database is currently available for the genes associated with ROS. Methods We present ROS- and diabetes-related targets (genes/proteins) collected from the biomedical literature through a text mining technology. A web-based literature mining tool, SciMiner, was applied to 1,154 biomedical papers indexed with diabetes and ROS by PubMed to identify relevant targets. Over-represented targets in the ROS-diabetes literature were obtained through comparisons against randomly selected literature. The expression levels of nine genes, selected from the top ranked ROS-diabetes set, were measured in the dorsal root ganglia (DRG) of diabetic and non-diabetic DBA/2J mice in order to evaluate the biological relevance of literature-derived targets in the pathogenesis of diabetic neuropathy. Results SciMiner identified 1,026 ROS- and diabetes-related targets from the 1,154 biomedical papers (http://jdrf.neurology.med.umich.edu/ROSDiabetes/). Fifty-three targets were significantly over-represented in the ROS-diabetes literature compared to randomly selected literature. These over-represented targets included well-known members of the oxidative stress response including catalase, the NADPH oxidase family, and the superoxide dismutase family of proteins. Eight of the nine selected genes exhibited significant differential expression between diabetic and non-diabetic mice. For six genes, the direction of expression change in diabetes paralleled enhanced oxidative stress in the DRG. Conclusions Literature mining compiled ROS-diabetes related targets from the biomedical literature and led us to evaluate the biological relevance of selected targets in the pathogenesis of diabetic neuropathy.http://deepblue.lib.umich.edu/bitstream/2027.42/78315/1/1755-8794-3-49.xmlhttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/2/1755-8794-3-49-S7.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/3/1755-8794-3-49-S10.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/4/1755-8794-3-49-S8.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/5/1755-8794-3-49-S3.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/6/1755-8794-3-49-S1.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/7/1755-8794-3-49-S4.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/8/1755-8794-3-49-S2.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/9/1755-8794-3-49-S12.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/10/1755-8794-3-49-S11.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/11/1755-8794-3-49-S9.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/12/1755-8794-3-49-S5.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/13/1755-8794-3-49-S6.XLShttp://deepblue.lib.umich.edu/bitstream/2027.42/78315/14/1755-8794-3-49.pdfPeer Reviewe

Daf-2 Signaling Modifies Mutant SOD1 Toxicity in C. elegans

The DAF-2 Insulin/IGF-1 signaling (IIS) pathway is a strong modifier of Caenorhabditis elegans longevity and healthspan. As aging is the greatest risk factor for developing neurodegenerative diseases such as Amyotrophic Lateral Sclerosis (ALS), we were interested in determining if DAF-2 signaling modifies disease pathology in mutant superoxide dismutase 1 (SOD1) expressing C. elegans. Worms with pan-neuronal G85R SOD1 expression demonstrate significantly impaired locomotion as compared to WT SOD1 expressing controls and they develop insoluble SOD1 aggregates. Reductions in DAF-2 signaling, either through a hypomorphic allele or neuronally targeted RNAi, decreases the abundance of aggregated SOD1 and results in improved locomotion in a DAF-16 dependant manner. These results suggest that manipulation of the DAF-2 Insulin/IGF-1 signaling pathway may have therapeutic potential for the treatment of ALS

Different Human Copper-Zinc Superoxide Dismutase Mutants, SOD1G93A and SOD1H46R, Exert Distinct Harmful Effects on Gross Phenotype in Mice

Amyotrophic lateral sclerosis (ALS) is a heterogeneous group of fatal neurodegenerative diseases characterized by a selective loss of motor neurons in the brain and spinal cord. Creation of transgenic mice expressing mutant Cu/Zn superoxide dismutase (SOD1), as ALS models, has made an enormous impact on progress of the ALS studies. Recently, it has been recognized that genetic background and gender affect many physiological and pathological phenotypes. However, no systematic studies focusing on such effects using ALS models other than SOD1G93A mice have been conducted. To clarify the effects of genetic background and gender on gross phenotypes among different ALS models, we here conducted a comparative analysis of growth curves and lifespans using congenic lines of SOD1G93A and SOD1H46R mice on two different genetic backgrounds; C57BL/6N (B6) and FVB/N (FVB). Copy number of the transgene and their expression between SOD1G93A and SOD1H46R lines were comparable. B6 congenic mutant SOD1 transgenic lines irrespective of their mutation and gender differences lived longer than corresponding FVB lines. Notably, the G93A mutation caused severer disease phenotypes than did the H46R mutation, where SOD1G93A mice, particularly on a FVB background, showed more extensive body weight loss and earlier death. Gender effect on survival also solely emerged in FVB congenic SOD1G93A mice. Conversely, consistent with our previous study using B6 lines, lack of Als2, a murine homolog for the recessive juvenile ALS causative gene, in FVB congenic SOD1H46R, but not SOD1G93A, mice resulted in an earlier death, implying a genetic background-independent but mutation-dependent phenotypic modification. These results indicate that SOD1G93A- and SOD1H46R-mediated toxicity and their associated pathogenic pathways are not identical. Further, distinctive injurious effects resulted from different SOD1 mutations, which are associated with genetic background and/or gender, suggests the presence of several genetic modifiers of disease expression in the mouse genome

Drosophila Carrying Pex3 or Pex16 Mutations Are Models of Zellweger Syndrome That Reflect Its Symptoms Associated with the Absence of Peroxisomes

The peroxisome biogenesis disorders (PBDs) are currently difficult-to-treat multiple-organ dysfunction disorders that result from the defective biogenesis of peroxisomes. Genes encoding Peroxins, which are required for peroxisome biogenesis or functions, are known causative genes of PBDs. The human peroxin genes PEX3 or PEX16 are required for peroxisomal membrane protein targeting, and their mutations cause Zellweger syndrome, a class of PBDs. Lack of understanding about the pathogenesis of Zellweger syndrome has hindered the development of effective treatments. Here, we developed potential Drosophila models for Zellweger syndrome, in which the Drosophila pex3 or pex16 gene was disrupted. As found in Zellweger syndrome patients, peroxisomes were not observed in the homozygous Drosophila pex3 mutant, which was larval lethal. However, the pex16 homozygote lacking its maternal contribution was viable and still maintained a small number of peroxisome-like granules, even though PEX16 is essential for the biosynthesis of peroxisomes in humans. These results suggest that the requirements for pex3 and pex16 in peroxisome biosynthesis in Drosophila are different, and the role of PEX16 orthologs may have diverged between mammals and Drosophila. The phenotypes of our Zellweger syndrome model flies, such as larval lethality in pex3, and reduced size, shortened longevity, locomotion defects, and abnormal lipid metabolisms in pex16, were reminiscent of symptoms of this disorder, although the Drosophila pex16 mutant does not recapitulate the infant death of Zellweger syndrome. Furthermore, pex16 mutants showed male-specific sterility that resulted from the arrest of spermatocyte maturation. pex16 expressed in somatic cyst cells but not germline cells had an essential role in the maturation of male germline cells, suggesting that peroxisome-dependent signals in somatic cyst cells could contribute to the progression of male germ-cell maturation. These potential Drosophila models for Zellweger syndrome should contribute to our understanding of its pathology

Connexin43 Modulates Cell Polarity and Directional Cell Migration by Regulating Microtubule Dynamics

Knockout mice deficient in the gap junction gene connexin43 exhibit developmental anomalies associated with abnormal neural crest, primordial germ cell, and proepicardial cell migration. These migration defects are due to a loss of directional cell movement, and are associated with abnormal actin stress fiber organization and a loss of polarized cell morphology. To elucidate the mechanism by which Cx43 regulates cell polarity, we used a wound closure assays with mouse embryonic fibroblasts (MEFs) to examine polarized cell morphology and directional cell movement. Studies using embryonic fibroblasts from Cx43 knockout (Cx43KO) mice showed Cx43 deficiency caused cell polarity defects as characterized by a failure of the Golgi apparatus and the microtubule organizing center to reorient with the direction of wound closure. Actin stress fibers at the wound edge also failed to appropriately align, and stabilized microtubule (Glu-tubulin) levels were markedly reduced. Forced expression of Cx43 with deletion of its tubulin-binding domain (Cx43dT) in both wildtype MEFs and neural crest cell explants recapitulated the cell migration defects seen in Cx43KO cells. However, forced expression of Cx43 with point mutation causing gap junction channel closure had no effect on cell motility. TIRF imaging revealed increased microtubule instability in Cx43KO cells, and microtubule targeting of membrane localized Cx43 was reduced with expression of Cx43dT construct in wildtype cells. Together, these findings suggest the essential role of Cx43 gap junctions in development is mediated by regulation of the tubulin cytoskeleton and cell polarity by Cx43 via a nonchannel function

Zebrafish Endzone Regulates Neural Crest-Derived Chromatophore Differentiation and Morphology

The development of neural crest-derived pigment cells has been studied extensively as a model for cellular differentiation, disease and environmental adaptation. Neural crest-derived chromatophores in the zebrafish (Danio rerio) consist of three types: melanophores, xanthophores and iridiphores. We have identified the zebrafish mutant endzone (enz), that was isolated in a screen for mutants with neural crest development phenotypes, based on an abnormal melanophore pattern. We have found that although wild-type numbers of chromatophore precursors are generated in the first day of development and migrate normally in enz mutants, the numbers of all three chromatophore cell types that ultimately develop are reduced. Further, differentiated melanophores and xanthophores subsequently lose dendricity, and iridiphores are reduced in size. We demonstrate that enz function is required cell autonomously by melanophores and that the enz locus is located on chromosome 7. In addition, zebrafish enz appears to selectively regulate chromatophore development within the neural crest lineage since all other major derivatives develop normally. Our results suggest that enz is required relatively late in the development of all three embryonic chromatophore types and is normally necessary for terminal differentiation and the maintenance of cell size and morphology. Thus, although developmental regulation of different chromatophore sublineages in zebrafish is in part genetically distinct, enz provides an example of a common regulator of neural crest-derived chromatophore differentiation and morphology

FUS and TARDBP but Not SOD1 Interact in Genetic Models of Amyotrophic Lateral Sclerosis

Mutations in the SOD1 and TARDBP genes have been commonly identified in Amyotrophic Lateral Sclerosis (ALS). Recently, mutations in the Fused in sarcoma gene (FUS) were identified in familial (FALS) ALS cases and sporadic (SALS) patients. Similarly to TDP-43 (coded by TARDBP gene), FUS is an RNA binding protein. Using the zebrafish (Danio rerio), we examined the consequences of expressing human wild-type (WT) FUS and three ALS–related mutations, as well as their interactions with TARDBP and SOD1. Knockdown of zebrafish Fus yielded a motor phenotype that could be rescued upon co-expression of wild-type human FUS. In contrast, the two most frequent ALS–related FUS mutations, R521H and R521C, unlike S57Δ, failed to rescue the knockdown phenotype, indicating loss of function. The R521H mutation caused a toxic gain of function when expressed alone, similar to the phenotype observed upon knockdown of zebrafish Fus. This phenotype was not aggravated by co-expression of both mutant human TARDBP (G348C) and FUS (R521H) or by knockdown of both zebrafish Tardbp and Fus, consistent with a common pathogenic mechanism. We also observed that WT FUS rescued the Tardbp knockdown phenotype, but not vice versa, suggesting that TARDBP acts upstream of FUS in this pathway. In addition we observed that WT SOD1 failed to rescue the phenotype observed upon overexpression of mutant TARDBP or FUS or upon knockdown of Tardbp or Fus; similarly, WT TARDBP or FUS also failed to rescue the phenotype induced by mutant SOD1 (G93A). Finally, overexpression of mutant SOD1 exacerbated the motor phenotype caused by overexpression of mutant FUS. Together our results indicate that TARDBP and FUS act in a pathogenic pathway that is independent of SOD1

Mitochondrial function as a determinant of life span

Average human life expectancy has progressively increased over many decades largely due to improvements in nutrition, vaccination, antimicrobial agents, and effective treatment/prevention of cardiovascular disease, cancer, etc. Maximal life span, in contrast, has changed very little. Caloric restriction (CR) increases maximal life span in many species, in concert with improvements in mitochondrial function. These effects have yet to be demonstrated in humans, and the duration and level of CR required to extend life span in animals is not realistic in humans. Physical activity (voluntary exercise) continues to hold much promise for increasing healthy life expectancy in humans, but remains to show any impact to increase maximal life span. However, longevity in Caenorhabditis elegans is related to activity levels, possibly through maintenance of mitochondrial function throughout the life span. In humans, we reported a progressive decline in muscle mitochondrial DNA abundance and protein synthesis with age. Other investigators also noted age-related declines in muscle mitochondrial function, which are related to peak oxygen uptake. Long-term aerobic exercise largely prevented age-related declines in mitochondrial DNA abundance and function in humans and may increase spontaneous activity levels in mice. Notwithstanding, the impact of aerobic exercise and activity levels on maximal life span is uncertain. It is proposed that age-related declines in mitochondrial content and function not only affect physical function, but also play a major role in regulation of life span. Regular aerobic exercise and prevention of adiposity by healthy diet may increase healthy life expectancy and prolong life span through beneficial effects at the level of the mitochondrion