Effects of DISC1 on Alzheimer's disease cell models assessed by iTRAQ proteomic analysis

Alzheimer’s disease (AD) is a form of neurodegenerative disease in the elderly with no cure at present. In a previous study, we found that the scaffold protein DISC1 is downregulated in the AD brains, and ectopic expression of DISC1 can delay the progression of AD by protecting synaptic plasticity and down-regulating BACE1. However, the underlying mechanisms remain not to be elucidated. In the present study, we compared the proteomes of normal and DISC1high AD cells expressing the amyloid precursor protein (APP) using isobaric tag for relative and absolute quantitation (iTRAQ) and mass spectrometry (MS). The differentially expressed proteins (DEPs) were identified, and the protein-protein interaction (PPI) network was constructed to identify the interacting partners of DISC1. Based on the interaction scores, NDE1, GRM3, PTGER3 and KATNA1 were identified as functionally or physically related to DISC1, and may therefore regulate AD development. The DEPs were functionally annotated by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases with the DAVID software, and the eggNOG database was used to determine their evolutionary relationships. The DEPs were significantly enriched in microtubules and mitochondria-related pathways. Gene set enrichment analysis (GSEA) was performed to identify genes and pathways that are activated when DISC1 is overexpressed. Our findings provide novel insights into the regulatory mechanisms underlying DISC1 function in AD.

207 Satellite Cells Participate in the wnt/ca2+ Pathway Controlled Porcine Myofiber Determination

The type of myofiber is important for porcine meat quality. Meanwhile, the nt/Ca2+ pathway has been showed multiple roles in skeletal muscle formation; however, the distinct mechanism is still unclear. In this study, the weaned piglets and satellite cells were designed into the control group, lysine deficiency group and lysine rescue group to investigate the function of Wnt/Ca2+ pathway in governing skeletal muscle typing. After we confirm the growth of weaned piglets was controlled by lysine, the isobaric tag for relative and absolute quantification (iTRAQ) analysis of skeletal muscle detected that Wnt/Ca2+ pathway was involved in the transition of fast and slow fiber. Then, we found the ratio of type I myofiber in Semimembranous (fast muscle) was significantly increased after lysine deficiency (P < 0.05), and decreased by lysine rescue (P < 0.05). In contrast, the ratio of type I myofiber in Semitendinous muscle (slow muscle) was significantly decreased in the lysine deficiency group, and increased in the lysine rescue group (P < 0.05). Furthermore, the Wnt/Ca2+ pathway was significantly increased in Semimembranous muscle, while decreased in Semitendinous muscle with lysine deficiency, and this phenomenon was inversed after lysine rescue (P < 0.05). Meanwhile, the Wnt/Ca2+ pathway was stronger in satellite cells isolated from Semitendinous muscle (StSCs) than that of Semimembranous satellite cell (SmSCs) (P < 0.05). And we also found the StSCs enter in differentiation is more easily than SmSCs (P < 0.05). Besides, the ratio of type I myofiber originated from StSCs showed greater than StSCs (P < 0.05). In summary, we conclude that satellite cells participate in the Wnt/Ca2+ pathway controlled porcine myofiber determination.

Aging Induces Hepatic Oxidative Stress and Nuclear Proteomic Remodeling in Liver from Wistar Rats

Aging is a continuous, universal, and irreversible process that determines progressive loss of adaptability. The liver is a critical organ that supports digestion, metabolism, immunity, detoxification, vitamin storage, and hormone signaling. Nevertheless, the relationship between aging and the development of liver diseases remains elusive. In fact, although prolonged fasting in adult rodents and humans delays the onset of the disease and increases longevity, whether prolonged fasting could exert adverse effects in old organisms remains incompletely understood. In this work, we aimed to characterize the oxidative stress and nuclear proteome in the liver of 3-month- and 24-month-old male Wistar rats upon 36 h of fasting and its adaptation in response to 30 min of refeeding. To this end, we analyzed the hepatic lipid peroxidation levels (TBARS) and the expression levels of genes associated with fat metabolism and oxidative stress during aging. In addition, to gain a better insight into the molecular and cellular processes that characterize the liver of old rats, the hepatic nuclear proteome was also evaluated by isobaric tag quantitation (iTRAQ) mass spectrometry-based proteomics. In old rats, aging combined with prolonged fasting had great impact on lipid peroxidation in the liver that was associated with a marked downregulation of antioxidant genes (Sod2, Fmo3, and Cyp2C11) compared to young rats. Besides, our proteomic study revealed that RNA splicing is the hepatic nuclear biological process markedly affected by aging and this modification persists upon refeeding. Our results suggest that aged-induced changes in the nuclear proteome could affect processes associated with the adaptative response to refeeding after prolonged fasting, such as those involved in the defense against oxidative stress.

Morphological observation and protein expression of fertile and abortive ovules in Castanea mollissima

Chinese chestnuts (Castanea mollissima Blume.) contain 12–18 ovules in one ovary, but only one ovule develops into a seed, indicating a high ovule abortion rate. In this study, the Chinese chestnut ‘Huaihuang’ was used to explore the possible mechanisms of ovule abortion with respect to morphology and proteomics. The morphology and microstructure of abortive ovules were found to be considerably different from those of fertile ovules at 20 days after anthesis (20 DAA). The fertile ovules had completely formed tissues, such as the embryo sac, embryo and endosperm. By contrast, in the abortive ovules, there were no embryo sacs, and wide spaces between the integuments were observed, with few nucelli. Fluorescence labelling of the nuclei and transmission electron microscopy (TEM) observations showed that cells of abortive ovules were abnormally shaped and had thickened cell walls, folded cell membranes, condensed cytoplasm, ruptured nuclear membranes, degraded nucleoli and reduced mitochondria. The iTRAQ (isobaric tag for relative and absolute quantitation) results showed that in the abortive ovules, low levels of soluble protein with small molecular weights were found, and most of differently expressed proteins (DEPs) were related to protein synthesis, accumulation of active oxygen free radical, energy synthesis and so on. These DEPs might be associated with abnormal ovules formation.

Proteomic Analysis of Red Ginseng on Prolonging the Life Span of Male Drosophila melanogaster

Ginseng (Panax ginseng C. A. Mey.) is a traditional medicine that has been utilized for over 2000 years in Asia and shows varied pharmacological effects. Red ginseng (RG) is steamed and dried ginseng root and is considered to be more effective. Heating inactivates its catabolic enzymes and increases the activities of RG, which can improve the immune system, alleviate fatigue, and has anti-inflammatory effects and antioxidant activity. In addition, RG has a good anti-aging effect, but its mechanism is unclear. Senescence, a side-effect of normal developmental and metabolic processes, is a gradual decline in physiological integrity and function of the body. Senescence is usually associated with a variety of diseases, including neurodegenerative diseases and diabetes. Research on anti-aging and the prolongation of life span has always been a focus topic. In this study, we investigated the molecular mechanism of RG that results in prolonged the life span for male Drosophila melanogaster. Isobaric tag for relative and absolute quantitation (iTRAQ) was used to identify protein changes in an old male D. melanogaster treated with RG. The differential proteins were verified by qRT-PCR and western blotting. The results showed that 12.5 mg/ml RG prolonged its life span significantly. iTRAQ results showed that, compared to the control group, 32 upregulated proteins and 62 downregulated proteins displayed significantly differential expression in the RG group. In this study, we explored the pathways that RG may participate in that extend the life span of D. melanogaster, and the results showed that the PI3K/AKT/FoxO pathway was involved. In addition, 4E-BP increased and participated in the regulation of life span.

Troxerutin-mediated C9 inhibition is a disease-modifying treatment for inflammatory arthritis.

Troxerutin (TXR) is a phytochemical reported to possess anti-inflammatory and hepatoprotective effects. In this study, we aimed to exploit anti-arthritic properties of TXR using an adjuvant induced arthritic (AIA) rat model. AIA induced rats showed highest arthritis score at disease onset and by oral administration of TXR (50, 100, 200 mg/kg body weight), reduced to basal level in a dose dependent manner. Isobaric tag for relative and absolute quantitative (iTRAQ) proteomics tool was employed to identify deregulated joint homogenate proteins in AIA and TXR treated rats to decipher probable mechanism of the TXR action in arthritis. iTRAQ analysis identified a set of 434 joint homogenate proteins with 65 deregulated proteins (log2 case/control ≥ 1.5) in AIA. Expressions of a set of important proteins (AAT, T-kininogen, vimentin, desmin, and nucleophosmin) that could classify AIA from healthy were validated using Western blot analysis. Western blot data corroborated proteomics findings. In silico protein-protein interaction study of joint homogenate proteome revealed that complement component 9 (C9), the major building blocks of the membrane attack complex (MAC) responsible for sterile inflammation, gets perturbed in AIA. Our dosimetry study suggests that a TXR dose of 200 mg/kg body weight for 15 days is sufficient to bring the arthritis score to basal levels in AIA rats. We have shown the importance of TXR as an anti-arthritis agent in AIA model and after additional investigation its arthritis ameliorating properties could be exploited for clinical usability.

Proteomic Profiling of the Liver, Hepatic Lymph Nodes, and Spleen of Buffaloes Infected with Fasciola gigantica

In the present study, we used an isobaric tag for relative and absolute quantitation (iTRAQ) proteomics technology to characterize the differentially expressed proteins (DEPs) in the liver, hepatic lymph nodes (hLNs), and spleen of buffaloes infected with Fasciola gigantica (F. gigantica). We also used the parallel reaction monitoring (PRM) method to verify the expression levels of the DEPs in the three infected tissues. At three days post-infection (dpi), 225, 1821, and 364 DEPs were detected in the liver, hLNs, and spleen, respectively. At 42 dpi, 384, 252, and 214 DEPs were detected in the liver, hLNs, and spleen, respectively. At 70 dpi, 125, 829, and 247 DEPs were detected in the liver, hLNs, and spleen, respectively. Downregulation of metabolism was prominent in infected livers at all time points, and upregulation of immune responses was marked in the hLNs during early infection (three dpi); however, no changes in the immune response were detected at the late stages of infection (42 and 70 dpi). Compared to the hLNs, there was no significant upregulation in the levels of immune responses in the infected spleen. All the identified DEPs were used to predict the subcellular localization of the proteins, which were related to extracellular space and membrane and were involved in host immune responses. Further PRM analysis confirmed the expression of 18 proteins. These data provide the first simultaneous proteomic profiles of multiple organs of buffaloes experimentally infected with F. gigantica.