Genome-wide identification of Bcl-3 and P50 target genes in disuse muscle atrophy

Thesis (Ph.D.)--Boston UniversityNF-κB plays a central role in regulating skeletal muscle atrophy. We previously showed that a NF-κB transcription factor p50 and its transcriptional coactivator Bcl-3 are both required for inducing muscular disuse atrophy due to hind limb unloading in a mouse model. However, less is known about the molecular mechanisms of the atrophy-resistant phenotype seen in nfkb1^-1- (lack of p50 protein) and bcl-3^-1- mice. The purpose of this study is to identify genes that are targets of p50 and Bcl-3 in unloading atrophy at a genome-wide level. Global gene expressions of plantar flexor muscles was first measured from wildtype, nfkb1^-1- and bcl-3^-1- mice with or without 6 days of hind limb unloading. Genes that were upregulated in wild-type mice due to unloading but not in the knockout mice were considered p50 or Bcl-3 direct or indirect target genes. 185 p50 and 240 Bcl-3 target genes were identified in disuse atrophy, and most of these genes encoded proteins involved in proteolysis and transcriptional regulation. All p50 target genes were also identified as Bcl-3 targets. Direct Bcl-3 binding targets in unloading atrophy were identified using ChIP-sequencing. In atrophied muscles there was an increase in Bcl-3 binding to the promoter regions of genes encoding E3 ligases, N-end rule proteins, kinase and glycolysis enzymes. By studying the expression changes of Bcl-3 binding targets, a Bcl-3 regulated gene network that is responsible for processes underlying unloading atrophy was mapped. One Bcl-3 direct target in unloading atrophy, Muscle specific Ring finger protein 1 (MuRF1), was studied in detail by qPCR, ChIP-seq, and MuRF1 promoter-reporter assays. These results provided the first direct evidence that the p50 and Bcl-3 bind to NF-κB binding sites to transactivate MuRF1 during muscle atrophy. Taken together, our data support Bcl-3 as a global regulator of genes involved in the proteolysis and the change in energy metabolism that are essential components of muscle atrophy due to disuse

The ChIP-seq-defined networks of Bcl-3 gene binding support its required role in skeletal muscle atrophy

NF-kappaB transcriptional activation is required for skeletal muscle disuse atrophy. We are continuing to study how the activation of NF-kB regulates the genes that encode the protein products that cause atrophy. Using ChIP-sequencing we found that Bcl-3, an NF-kB transcriptional activator required for atrophy, binds to the promoters of a number of genes whose collective function describes two major aspects of muscle wasting. By means of bioinformatics analysis of ChIP-sequencing data we found Bcl-3 to be directing transcription networks of proteolysis and energy metabolism. The proteolytic arm of the Bcl-3 networks includes many E3 ligases associated with proteasomal protein degradation, including that of the N-end rule pathway. The metabolic arm appears to be involved in organizing the change from oxidative phosphorylation to glycolysis in atrophying muscle. For one gene, MuRF1, ChIP-sequencing data identified the location of Bcl-3 and p50 binding in the promoter region which directed the creation of deletant and base-substitution mutations of MuRF1 promoter constructs to determine the effect on gene transcription. The results provide the first direct confirmation that the NF-kB binding site is involved in the muscle unloading regulation of MuRF1. Finally, we have combined the ChIP-sequencing results with gene expression microarray data from unloaded muscle to map several direct targets of Bcl-3 that are transcription factors whose own targets describe a set of indirect targets for NF-kB in atrophy. ChIP-sequencing provides the first molecular explanation for the finding that Bcl3 knockout mice are resistant to disuse muscle atrophy. Mapping the transcriptional regulation of muscle atrophy requires an unbiased analysis of the whole genome, which we show is now possible with ChIP-sequencing.R01 AR041705 - NIAMS NIH HHS; R01 AR060217 - NIAMS NIH HHS; AR041705 - NIAMS NIH HHS; AR060217 - NIAMS NIH HH

RNA-seq and metabolomic analyses of Akt1-mediated muscle growth reveals regulation of regenerative pathways and changes in the muscle secretome

A schematic representation of gene expression changes in the oxidative phosphorylation pathway. (PDF 590 kb

Note on a Single-Machine Scheduling Problem with Sum of Processing Times Based Learning and Ready Times

In the recent 20 years, scheduling with learning effect has received considerable attention. However, considering the learning effect along with release time is limited. In light of these observations, in this paper, we investigate a single-machine problem with sum of processing times based learning and ready times where the objective is to minimize the makespan. For solving this problem, we build a branch-and-bound algorithm and a heuristic algorithm for the optimal solution and near-optimal solution, respectively. The computational experiments indicate that the branch-and-bound algorithm can perform well the problem instances up to 24 jobs in terms of CPU time and node numbers, and the average error percentage of the proposed heuristic algorithm is less than 0.5%

Treatment of Color Filter Wastewater by Fresnel Lens Enhanced Solar Photo-Fenton Process

Treatment of color filter wastewater using solar photo-Fenton process enhanced by high-concentrating Fresnel lens was investigated in this paper. Optimal reaction conditions based on response surface methodology (RSM) were established as under an initial pH of 5, a [H2O2]0/COD0 ratio of 1~1.35 and a [H2O2]0/[Fe2+]0 ratio of 15 for a reaction time of 60 min, which could reach a readily biodegradable level, that is, the biodegradability (BOD5/COD) of wastewater was more than 0.3. With the assistance of Fresnel lens, the solar photo-Fenton process increased the COD degradation rate and mineralization rate by a factor of 4.5 and 6.5, respectively. In addition, the microtoxicity (TU50) of wastewater was almost diminished after a 60 min of treatment, whereas the microtoxicity of treated wastewater without the assistance of Fresnel lens remained a TU50 value of 1.166. This could be mainly due to the concentrating effect of Fresnel lens for solar energy, resulting in an increase of 2~3 times of solar light intensity and a raising heat irradiation in terms of 15~30 °C of wastewater temperature. These results revealed that solar energy could be concentrated effectively by using Fresnel lens and showed a significant promoting effect on the photo-Fenton reaction for treating color filter wastewater

The study on SFLAB GanedenBC30 viability on baking products during storage

AbstractFor understanding Bacillus coagulans, GanedenBC30 was used in different ways to added in raw dough and examine their viability after baking. Eight different baking products: (1) chrysanthemum cookies, (2) egg pastry cakes, (3) mooncakes, (4) muffins, (5) polo breads, (6) soda cookies, (7) sponge cakes, and (8) toasts were made from 0.5% GanedenBC30 added to their dough in two ways: (a) flour powder or (b) egg yolk. Then the (a) pH value, (b) titratable acidity, (c) GanedenBC30 counts, and (d) viability GanedenBC30 of eight different baking products were determined after storing at 4oC for 0, 3, 6, 9, 12, 15 days, or 25oC for 0, 3, 6 days. The eight types of raw dough had relatively lower pH values and rise after baking. The titratable acidity of the eight types of dough was relatively higher, and declined after baking. However, the pH value and titratable acidity of the eight baking products remained the same after 9 days at 4oC. On the other hand, the GanedenBC30 counts in the eight baking products were less than their raw dough GanedenBC30 levels. For storage at both 4 and 25oC, the results show the GanedenBC30 viability of baking products decreased with storage days. The dough made by flour powder and baking showed higher GanedenBC30 viability than by egg yolk. GanedenBC30 are good candidates for baking product use, both in lactic acid production and probiotic preparations

Genome-wide identification of FoxO-dependent gene networks in skeletal muscle during C26 cancer cachexia

BACKGROUND: Evidence from cachectic cancer patients and animal models of cancer cachexia supports the involvement of Forkhead box O (FoxO) transcription factors in driving cancer-induced skeletal muscle wasting. However, the genome-wide gene networks and associated biological processes regulated by FoxO during cancer cachexia are unknown. We hypothesize that FoxO is a central upstream regulator of diverse gene networks in skeletal muscle during cancer that may act coordinately to promote the wasting phenotype. METHODS: To inhibit endogenous FoxO DNA-binding, we transduced limb and diaphragm muscles of mice with AAV9 containing the cDNA for a dominant negative (d.n.) FoxO protein (or GFP control). The d.n.FoxO construct consists of only the FoxO3a DNA-binding domain that is highly homologous to that of FoxO1 and FoxO4, and which outcompetes and blocks endogenous FoxO DNA binding. Mice were subsequently inoculated with Colon-26 (C26) cells and muscles harvested 26 days later. RESULTS: Blocking FoxO prevented C26-induced muscle fiber atrophy of both locomotor muscles and the diaphragm and significantly spared force deficits. This sparing of muscle size and function was associated with the differential regulation of 543 transcripts (out of 2,093) which changed in response to C26. Bioinformatics analysis of upregulated gene transcripts that required FoxO revealed enrichment of the proteasome, AP-1 and IL-6 pathways, and included several atrophy-related transcription factors, including Stat3, Fos, and Cebpb. FoxO was also necessary for the cancer-induced downregulation of several gene transcripts that were enriched for extracellular matrix and sarcomere protein-encoding genes. We validated these findings in limb muscles and the diaphragm through qRT-PCR, and further demonstrate that FoxO1 and/or FoxO3a are sufficient to increase Stat3, Fos, Cebpb, and the C/EBPβ target gene, Ubr2. Analysis of the Cebpb proximal promoter revealed two bona fide FoxO binding elements, which we further establish are necessary for Cebpb promoter activation in response to IL-6, a predominant cytokine in the C26 cancer model. CONCLUSIONS: These findings provide new evidence that FoxO-dependent transcription is a central node controlling diverse gene networks in skeletal muscle during cancer cachexia, and identifies novel candidate genes and networks for further investigation as causative factors in cancer-induced wasting.R01 AR060217 - NIAMS NIH HHS; R01 AR060209 - NIAMS NIH HHS; T32 HD043730 - NICHD NIH HHS; R00 HL098453 - NHLBI NIH HHS; R00HL098453 - NHLBI NIH HHS; R01AR060209 - NIAMS NIH HHS; R01AR060217 - NIAMS NIH HH

Effect on Spasticity After Performance of Dynamic-Repeated-Passive Ankle Joint Motion Exercise in Chronic Stroke Patients

Spasticity associated with abnormal muscle tone is a common motor disorder following stroke, and the spastic ankle may affect ambulatory function. The purpose of this study was to investigate the short-term effect of dynamic-repeated-passive ankle movements with weight loading on ambulatory function and spastic hypertonia of chronic stroke patients. In this study, 12 chronic stroke patients with ankle spasticity and inefficient ambulatory ability were enrolled. Stretching of the plantar-flexors of the ankle in the standing position for 15 minutes was performed passively by a constant-speed and electrically powered device. The following evaluations were done before and immediately after the dynamic-repeated-passive ankle movements. Spastic hypertonia was assessed by the Modified Ashworth Scale (MAS; range, 0–4), Achilles tendon reflexes test (DTR; range, 0–4), and ankle clonus (range, 0–5). Improvement in ambulatory ability was determined by the timed up-and-go test (TUG), the 10-minute walking test, and cadence (steps/minute). In addition, subjective experience of the influence of ankle spasticity on ambulation was scored by visual analog scale (VAS). Subjective satisfaction with the therapeutic effect of spasticity reduction was evaluated by a five-point questionnaire (1 = very poor, 2 = poor, 3 = acceptable, 4 = good, 5 = very good). By comparison of the results before and after intervention, these 12 chronic stroke patients presented significant reduction in MAS and VAS for ankle spasticity, the time for TUG and 10-minute walking speed (p < 0.01). The cadence also increased significantly (p < 0.05). In addition, subjective satisfaction with the short-term therapeutic effect was mainly good (ranging from acceptable to very good). In conclusion, 15 minutes of dynamic-repeated-passive ankle joint motion exercise with weight loading in the standing position by this simple constant-speed machine is effective in reducing ankle spasticity and improving ambulatory ability