Module-based multiscale simulation of angiogenesis in skeletal muscle

Background  

Mathematical modeling of angiogenesis has been gaining momentum as a means to shed new light on the biological complexity underlying blood vessel growth. A variety of computational models have been developed, each focusing on different aspects of the angiogenesis process and occurring at different biological scales, ranging from the molecular to the tissue levels. Integration of models at different scales is a challenging and currently unsolved problem.     

Different population histories of the Mundari- and Mon-Khmer-speaking Austro-Asiatic tribes inferred from the mtDNA 9-bp deletion/insertion polymorphism in Indian populations

Length variation in the human mtDNA intergenic region between the cytochrome oxidase II (COII) and tRNA lysine (tRNA^lys) genes has been widely studied in world populations. Specifically, Austronesian populations of the Pacific and Austro-Asiatic populations of southeast Asia most frequently carry the 9-bp deletion in that region implying their shared common ancestry in haplogroup B. Furthermore, multiple independent origins of the 9-bp deletion at the background of other mtDNA haplogroups has been shown in populations of Africa, Europe, Australia, and India. We have analyzed 3293 Indian individuals belonging to 58 populations, representing different caste, tribal, and religious groups, for the length variation in the 9-bp motif. The 9-bp deletion (one copy) and insertion (three copies) alleles were observed in 2.51% (2.15% deletion and 0.36% insertion) of the individuals. The maximum frequency of the deletion (45.8%) was observed in the Nicobarese in association with the haplogroup B5a D-loop motif that is common throughout southeast Asia. The low polymorphism in the D-loop sequence of the Nicobarese B5a samples suggests their recent origin and a founder effect, probably involving migration from southeast Asia. Interestingly, none of the 302 (except one Munda sample, which has 9-bp insertion) from Mundari-speaking Austro-Asiatic populations from the Indian mainland showed the length polymorphism of the 9-bp motif, pointing either to their independent origin from the Mon-Khmeric-speaking Nicobarese or to an extensive admixture with neighboring Indo-European-speaking populations. Consistent with previous reports, the Indo-European and Dravidic populations of India showed low frequency of the 9-bp deletion/insertion. More than 18 independent origins of the deletion or insertion mutation could be inferred in the phylogenetic analysis of the D-loop sequences.     

Caprylic Triglyceride as a Novel Therapeutic Approach to Effectively Improve the Performance and Attenuate the Symptoms Due to the Motor Neuron Loss in ALS Disease

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder of motor neurons causing progressive muscle weakness, paralysis, and finally death. ALS patients suffer from asthenia and their progressive weakness negatively impacts quality of life, limiting their daily activities. They have impaired energy balance linked to lower activity of mitochondrial electron transport chain enzymes in ALS spinal cord, suggesting that improving mitochondrial function may present a therapeutic approach for ALS. When fed a ketogenic diet, the G93A ALS mouse shows a significant increase in serum ketones as well as a significantly slower progression of weakness and lower mortality rate. In this study, we treated SOD1-G93A mice with caprylic triglyceride, a medium chain triglyceride that is metabolized into ketone bodies and can serve as an alternate energy substrate for neuronal metabolism. Treatment with caprylic triglyceride attenuated progression of weakness and protected spinal cord motor neuron loss in SOD1-G93A transgenic animals, significantly improving their performance even though there was no significant benefit regarding the survival of the ALS transgenic animals. We found that caprylic triglyceride significantly promoted the mitochondrial oxygen consumption rate in vivo. Our results demonstrated that caprylic triglyceride alleviates ALS-type motor impairment through restoration of energy metabolism in SOD1-G93A ALS mice, especially during the overt stage of the disease. These data indicate the feasibility of using caprylic acid as an easily administered treatment with a high impact on the quality of life of ALS patients.     

Lack of Cytochrome c in Mouse Fibroblasts Disrupts Assembly/Stability of Respiratory Complexes I and IV

Cytochrome c (cyt c) is a heme-containing protein that participates in electron transport in the respiratory chain and as a signaling molecule in the apoptotic cascade. Here we addressed the effect of removing mammalian cyt c on the integrity of the respiratory complexes in mammalian cells. Mitochondria from cyt c knockout mouse cells lacked fully assembled complexes I and IV and had reduced levels of complex III. A redox-deficient mutant of cyt c was unable to rescue the levels of complexes I and IV. We found that cyt c is associated with both complex IV and respiratory supercomplexes, providing a potential mechanism for the requirement for cyt c in the assembly/stability of complex IV.The mitochondrial electron transport chain consists of four multisubunit complexes, namely, NADH-ubiquinone oxidoreductase (complex I),² succinate-ubiquinone oxidoreductase (complex II), ubiquinone-cytochrome c oxidoreductase (complex III), and cytochrome c oxidase (complex IV, COX). Cytochrome c (cyt c) shuttles electrons from oxidative phosphorylation complex III to complex IV. Electrons are transferred from reduced cyt c sequentially to the Cu_A site, heme a, heme a₃, and Cu_B binuclear center in the complex IV before being finally transferred to molecular oxygen to generate water (1). Respiratory complexes are assembled into supercomplexes (also called respirasomes). These contain complex I bound to dimeric complex III and a variable copy number of complex IV (2).In Saccharomyces cerevisiae, cyt c is encoded by two genes: CYC1 and CYC7. Mutagenesis studies in yeast have shown that cyt c is required for the assembly of COX (3, 4). In yeast lacking both the cyt c genes (CYC1 and CYC7), COX assembly was absent. It was also shown that cyt c is only structurally required for COX assembly, because a catalytic mutant of cyt c (W65S) was sufficient to bring about near normal levels of COX. However, because yeast lacks complex I, they could not analyze the role of cyt c in the assembly/stability of complex I. Mammals possess two different isoforms of cyt c encoded on different chromosomes: the somatic (cyt cS)- and testis (cyt cT)-specific isoforms. In mouse, the cDNAs bear 74% homology, whereas the proteins possess 86% identity with most dissimilarity in the C terminus.Cardiolipin (CL) is an anionic phospholipid present almost exclusively in the mitochondrial membranes and constitutes 25% of its total phospholipids (5). Work from several laboratories showed that CL is essential for the membrane anchorage of the respiratory supercomplexes. CL has two main roles in the mitochondrial structure and function, namely, stabilization of mitochondrial membranes and specific interactions with proteins. CL deficiency results in inefficient energy transformation by oxidative phosphorylation, swelling of mitochondria, decreased ATP/oxygen ratio, and reduced membrane potential (6, 7). In accordance, in S. cerevisiae lacking CL synthase, the supercomplex comprising complexes III and IV is unstable (8). Assembly mutants of COX had significantly reduced CL synthase activity, whereas assembly mutants of respiratory complex III and complex V showed less inhibition (9). Subsequently, the proton gradient across the inner mitochondrial membrane was found to be important for CL formation and that CL synthase was stimulated by alkaline pH at the matrix side (10). In this study, we investigated the role of cyt c depletion on CL levels by examining its content and composition in cyt c null cells.Here we aimed to answer the following questions: What is the role of cyt c in the assembly and maintenance of the different respiratory complexes in mammals? Are there changes in the content/composition of lipids in the cyt c-ablated cells? Analysis of mouse fibroblasts revealed that cyt c is essential for the assembly/stability of COX, and a catalytically mutant form of cyt c cannot rescue the COX defect in the cyt c null cells. CL and triacylglycerols showed significant differences in the cyt c null cells, both in content and composition.     

Isolation and characterization of a transforming growth factor-beta Type II receptor cDNA from Xenopus laevis

Transforming Growth Factor-beta (TGF-beta) and their receptors have been characterized from many organisms. Two TGF-beta signaling receptors called Type I and II have been described for various ligands of the superfamily from organisms ranging from Drosophila to humans. In Xenopus laevis, TGF-beta2 and 5 have been reported and presumably, play important roles during early development. Several Type I and type II receptors for many ligands of the TGF-beta superfamily except TGF-beta type II receptor (TbetaIIR), have been characterized in Xenopus laevis. A chemical cross linking experiment using iodinated TGF-beta1 and -beta5, revealed four specific binding proteins on XTC cells. In order to understand the TGF-beta involvement during Xenopus development, a TGF-beta type II receptor (XTbetaIIR) has been isolated from a XTC cDNA library. XTbetaIIR was a partial cDNA lacking a portion of the signal peptide. The sequence analysis and homology comparison with the human TbetaIIR revealed 67% amino acid similarity in the extra cellular domain, 60% similarity in the transmembrane domain and 87% similarity in the cytoplasmic kinase domain, suggesting that XTbetaIIR is a putative TGF-beta type II receptor. In addition, the consensus amino acid motif for serine threonine receptor kinases was also present. Further, a dominant negative expression construct lacking the cytoplasmic kinase domain (engineered with the signal peptide from human TGF-beta type II receptor), was able to abolish TGF-beta mediated induction of a luciferase reporter plasmid, in a transient cell transfection assay. This substantiates the notion that XTbetaIIR cDNA can act as a receptor for TGF-beta. RT-PCR analysis using RNA isolated from various developmental stages of Xenopus laevis revealed expression of this gene in all the early stages of development and in the adult organs, except in stages 46/48.     

DNA damage in the presence of chemical genotoxic agents induce acetylation of H3K56 and H4K16 but not H3K9 in mammalian cells

Histone covalent modifications play a significant role in the regulation of chromatin structure and function during DNA damage. Hyperacetylation of histones is a DNA damage dependent post translational modification in yeast and mammals. Although acetylation of histones during DNA damage is well established, specific lysine residues that are acetylated is being understood very recently in mammals. Here, in the present study, acetylation of three different lysine residues Histone3Lysine 9 (H3K9), Histone3Lysine 56 (H3K56) and Histone4Lysine 16 (H4K16) were probed with specific antibodies in mammalian cell lines treated with genotoxic agents that induce replication stress or S-phase dependent double strand breaks. Immunoblotting results have shown that DNA damage associated with replication arrest induce acetylation of H3K56 and H4K16 but not H3K9 in mammals. Immunofluorescence experiments further confirmed that acetylated H3K56 and H4K16 form nuclear foci at the site of DNA double strand breaks. Colocalization of H3K56ac with γ H2AX and replication factor PCNA proved the existence of this modification at the site of DNA damage and its probable role in DNA damage repair. Put together, the present data suggests that acetylation of H3K56 and H4K16 are potent DNA damage dependent histone modifications but not H3K9 in mammals.     

Ferrous Iron Treatment of Soils Contaminated with Arsenic-Containing Wood-Preserving Solution

This article discusses the results of efforts to reclaim As-contaminated soil from a former timber-treating plant. The study site, commonly referred to as the Rocker Timber Framing site, is located along Silver Bow Creek approximately 7 miles west of the Butte Mining District, MT, USA. The plant operations resulted in contamination of the soils with a highly caustic solution containing 5% As (III). Contaminated soil resulted in the groundwater plumes that contained up to 25?mg L^?1 As, with As (V) being the predominant species. The objective of this study was to evaluate the effectiveness of Fe (II) treatment for remediation of As-contaminated soils. Laboratory-treatability studies were conducted on samples of saturated zone (AS1) and va-dose zone (AV1) soils. The AS1 soil was a mixture of coarse alluvium and potentially some mill tailings from adjacent mining operations. The AV1 soil consisted primarily of fill, including soil, construction debris, and timber fragments. Initial concentrations of total As in AS1 and AV1 soils were 683 and 4814?µg kg^?1, respectively. Water-soluble As concentrations were 15.4 and 554?µg L^?1, respectively, in a 20:1 solution to soil extract. Batch equilibration were performed by placing 10?g of soil into 20 vessels and adding increasing amounts of FeSO₄.7H₂O. Amendment increments were made as multiples of molar ratios of total As present in each soil. Treatability studies were run with and without a pH buffer of CaCO₃ (added at a 2:1 molar ratio to the FeSO₄.7H₂O treatment). Solution concentrations of As in the AS1 and AV1 soils (without CaCO₃) decreased from 554 to 15.4?µ L^?1 and 3802 to 0.64?µ L^?1, respectively, as the Fe:As molar ratios increased from 0 to 2, whereas for the AS1 soil the solution As concentration increased at the Fe:As molar ratios >2 and reverse trend was observed for the AV1 soils. The decrease in As solution concentration for the AS1 soil is attributable to the dramatic decrease in soil pH with increasing Fe:As molar ratios. In the case of soils treated with CaCO₃, the solution concentrations decreased from 564 to 0.65?µg L^?1 and 3790 to 0.79?µg L^?1 for the AS1 and AV1 soils, respectively,as the Fe:As molar ratios increased from 0 to 50. Generally, in both the soils, the CaCO₃-treated soil contained significantly more solution As compared with the non-CaCO₃-treated soil at the comparable Fe:As molar ratios. This is attributable to higher solution pH on CaCO₃ treatment. Our rapid engineering study indicates that treating both the soils with Fe:As molar ratio of 2 lowered the As water quality limit to <50?µL^?1, whereas treating the AS1 and AV1 soils with Fe:As molar ratio of 2 and 3, respectively, lowered the As water quality limit to ≤15?µg L^?1. The concentrations of the Cu and Zn were below the instrument detection limits for the AS1 and AV1 soils without CaCO₃ treatment. Sequential extraction of Fe-treated soils illustrated that As was relatively stable. Less than 1% of the As was extractable using a modified TCLP approach and <70% of the As was extractable using a harsh acid modified hydroxylamine hydrochloride extraction.     

Mouse models of oxidative phosphorylation defects: powerful tools to study the pathobiology of mitochondrial diseases

Defects in the oxidative phosphorylation system (OXPHOS) are responsible for a group of extremely heterogeneous and pleiotropic pathologies commonly known as mitochondrial diseases. Although many mutations have been found to be responsible for OXPHOS defects, their pathogenetic mechanisms are still poorly understood. An important contribution to investigate the in vivo function of several mitochondrial proteins and their role in mitochondrial dysfunction, has been provided by mouse models. Thanks to their genetic and physiologic similarity to humans, mouse models represent a powerful tool to investigate the impact of pathological mutations on metabolic pathways. In this review we discuss the main mouse models of mitochondrial disease developed, focusing on the ones that directly affect the OXPHOS system.     

p300-mediated Acetylation of Histone H3 Lysine 56 Functions in DNA Damage Response in Mammals

The packaging of newly replicated and repaired DNA into chromatin is crucial for the maintenance of genomic integrity. Acetylation of histone H3 core domain lysine 56 (H3K56ac) has been shown to play a crucial role in compaction of DNA into chromatin following replication and repair in Saccharomyces cerevisiae. However, the occurrence and function of such acetylation has not been reported in mammals. Here we show that H3K56 is acetylated and that this modification is regulated in a cell cycle-dependent manner in mammalian cells. We also demonstrate that the histone acetyltransferase p300 acetylates H3K56 in vitro and in vivo, whereas hSIRT2 and hSIRT3 deacetylate H3K56ac in vivo. Further we show that following DNA damage H3K56 acetylation levels increased, and acetylated H3K56, which is localized at the sites of DNA repair. It also colocalized with other proteins involved in DNA damage signaling pathways such as phospho-ATM, CHK2, and p53. Interestingly, analysis of occurrence of H3K56 acetylation using ChIP-on-chip revealed its genome-wide spread, affecting genes involved in several pathways that are implicated in tumorigenesis such as cell cycle, DNA damage response, DNA repair, and apoptosis.     

Peroxisome proliferator activator receptor gamma coactivator-1alpha (PGC-1α) improves motor performance and survival in a mouse model of amyotrophic lateral sclerosis

Background

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease that affects spinal cord and cortical motor neurons. An increasing amount of evidence suggests that mitochondrial dysfunction contributes to motor neuron death in ALS. Peroxisome proliferator-activated receptor gamma co-activator-1α (PGC-1α) is a principal regulator of mitochondrial biogenesis and oxidative metabolism.

Results

In this study, we examined whether PGC-1α plays a protective role in ALS by using a double transgenic mouse model where PGC-1α is over-expressed in an SOD1 transgenic mouse (TgSOD1-G93A/PGC-1α). Our results indicate that PGC-1α significantly improves motor function and survival of SOD1-G93A mice. The behavioral improvements were accompanied by reduced blood glucose level and by protection of motor neuron loss, restoration of mitochondrial electron transport chain activities and inhibition of stress signaling in the spinal cord.

Conclusion

Our results demonstrate that PGC-1α plays a beneficial role in a mouse model of ALS, suggesting that PGC-1α may be a potential therapeutic target for ALS therapy.