Comparative proteome analysis of splenic lymphocytes in long-term high-fat diet and dietary supplement with lipoic acid mice

The objective of this investigation was to explore possible molecular changes for role of a high-fat diet (HFD)-induced oxidative stress in splenic lymphocytes, and whether a dietary lipoic acid (LA) supplement could attenuate these changes. Male C57BL/6 mice were fed one of three diets 10 weeks and outcome measures centered on parameters of oxidative stress and lymphocytes apoptosis in spleen. Two-dimensional gel electrophoresis was used to compare the proteomes of splenic lymphocytes with three dietary groups. Differentially expressed spots whose expression altered over three fold were identified by MALDI-TOF MS. In this study, HFD resulted in oxidative stress in mice spleen, and significantly increased apoptotic percentage of splenic lymphocytes. Bioinformatic evaluation results of MALDI-TOF MS showed that 20 differentially expressed protein spots were known to be involved in many processes associated with cell function, such as cytoskeleton, energy metabolism and oxidative stress, signal transduction and cell defense. In conclusion, these results indicate that HFD-induced oxidative stress could lead to the functional decline of splenic lymphocytes, and LA supplement attenuates the alterations of protein expression to maintain the basic biological processes.     

Gene expression profiles in human cells submitted to genotoxic stress

Cell response to genotoxic agents is complex and involves the participation of different classes of genes (DNA repair, cell cycle control, signal transduction, apoptosis and oncogenesis). In this report, we present three approaches to document gene expression profiles, dealing with the evaluation of cellular responses to genotoxic agents (gamma-rays from 60Cobalt and cyclophosphamide). We used the method of cDNA arrays to analyze the differential gene expression profiles that were displayed by lymphocytes from radiation-exposed individuals, a human fibroblast cell line, and T lymphocytes from systemic lupus erythematosus (SLE) patients who were treated with cyclophosphamide. A preliminary analysis performed in lymphocytes from three radiation-workers showed that several induced genes can be associated with cell response to ionizing radiation: TRRAP (cell cycle regulation), Ligase IV (DNA repair), MAPK8IP1 and MAPK10 (signal transduction), RASSF2 (apoptosis induction/tumorigenesis), p53 (damage response/maintenance of genetic stability). The in vitro irradiated normal VH16 cell line (primary) showed a complex response to the genotoxic stress at the molecular level. Many apoptotic pathways were concomitantly induced. In addition, several genes involved in signaling and cell cycle arrest/control were significantly modulated after irradiation. Many genes involved in oxidative damage were also induced, indicating that this mechanism seems to be an important component of cell response. After treatment of the SLE patients with cyclophosphamide, 154 genes were differentially and significantly induced. Among them, we identified those associated with drug detoxification, cell cycle control, apoptosis, and tumor-suppressor. These findings indicate that at least two apoptotic pathways were induced after cyclophosphamide treatment. The induction of APAF1 and two genes coding for two subunits of cytochrome c supports a previous report showing increased apoptosis in lymphocytes from SLE patients. The present study provides new information on the molecular mechanism underlying the cell response to genotoxic stress, with relevance to basic and clinical research.     

Grape Seed and Skin Extract Prevents High-Fat Diet-Induced Brain Lipotoxicity in Rat

Obesity is related to an elevated risk of dementia and the physiologic mechanisms whereby fat adversely affects the brain are poorly understood. The present investigation analyzed the effect of a high fat diet (HFD) on brain steatosis and oxidative stress and the intracellular mediators involved in signal transduction, as well as the protection offered by grape seed and skin extract (GSSE). HFD induced ectopic deposition of cholesterol and phospholipid but not triglyceride. Moreover brain lipotoxicity is linked to an oxidative stress characterized by increased lipoperoxidation and carbonylation, inhibition of glutathione peroxidase and superoxide dismutase activities, depletion of manganese and a concomitant increase in ionizable calcium and acetylcholinesterase activity. Importantly GSSE alleviated all the deleterious effects of HFD treatment. Altogether our data indicated that HFD could find some potential application in the treatment of manganism and that GSSE should be used as a safe anti-lipotoxic agent in the prevention and treatment of fat-induced brain injury.     

Immune response gene 1 deficiency aggravates high fat diet-induced nonalcoholic fatty liver disease via promotion of redox-sensitive AKT suppression

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disorder worldwide. Immune response gene 1 (IRG1) catalyzes the production of bio-active itaconate, which is actively involved in the regulation of signal transduction. A recent study has found that the expression of IRG1 was significantly down-regulated in obesity-associated fatty liver, but the potential roles of IRG1 in the development NAFLD remain unclear. The present study found that genetic deletion of IRG1 aggravated high fat diet (HFD)-induced metabolic disturbance, including obesity, dyslipidemia and insulin resistance. In addition, HFD induced more severe liver steatosis and higher serum ALT and AST level in IRG1 KO mice, which were accompanied with altered expression of genes involved in lipid uptake, synthesis and catabolism. RNA-seq and immunoblot analysis indicated that deficiency of IRG1 is associated with suppressed activation of AKT, a master metabolic regulator. Mechanistically, IRG1/itaconate enhanced the antioxidative NRF2 pathway and prevented redox-sensitive suppression of AKT. Interestingly, supplementation with 4-octyl itaconate (4-OI), a cell-permeable derivate of itaconate, alleviated HFD-induced oxidative stress, AKT suppression and liver steatosis. Therefore, IRG1 probably functions as a protective regulator in the development of NAFLD and the cell-permeable 4-OI might have potential value for the pharmacological intervention of NAFLD.     

铝诱导植物程序性细胞死亡信号转导的研究进展

铝是制约酸性土壤上作物生产的主要因素。铝诱导氧化胁迫产生大量活性氧/一氧化氮,引起胞质钙超载,通过线粒体信号转导途径激发相关凋亡基因,从而引起细胞主动死亡,以减轻铝对植物的进一步毒害。本文综述了铝诱导程序性细胞死亡的信号分子、相关基因以及信号转导途径,对未来的研究方向提出了展望,为深入研究植物铝毒害机理和耐铝机制提供参考。     

CTRP9 knockout exaggerates lipotoxicity in cardiac myocytes and high‐fat diet‐induced cardiac hypertrophy through inhibiting the LKB1/AMPK pathway

CTRP9 has been reported to regulate lipid metabolism and exert cardioprotective effects, yet its role in high‐fat diet (HFD)‐induced cardiac lipotoxicity and the underlying mechanisms remain unclear. In the current study, we established HFD‐induced obesity model in wild‐type (WT) or CTRP9 knockout (CTRP9‐KO) mice and palmitate‐induced lipotoxicity model in neonatal rat cardiac myocytes (NRCMs) to investigate the effects of CTRP9 on cardiac lipotoxicity. Our results demonstrated that the HFD‐fed CTRP9‐KO mice accentuated cardiac hypertrophy, fibrosis, endoplasmic reticulum (ER) stress‐initiated apoptosis and oxidative stress compared with the HFD‐fed WT mice. In vitro, CTRP9 treatment markedly alleviated palmitate‐induced oxidative stress and ER stress‐induced apoptosis in NRCMs in a dose‐dependent manner. Phosphorylated AMPK at Thr172 was reduced, and phosphorylated mammalian target of rapamycin (mTOR) was strengthened in the heart of the HFD‐fed CTRP9‐KO mice compared with the HFD‐fed control mice. In vitro, AMPK inhibitor compound C significantly abolished the effects of CTRP9 on the inhibition of the apoptotic pathway in palmitate‐treated NRCMs. In a further mechanistic study, CTRP9 enhanced expression of phosphorylated LKB1 at Ser428 and promoted LKB1 cytoplasmic localization. Besides, silencing of LKB1 gene by lentivirus significantly prohibited activation of AMPK by CTRP9 and partially eliminated the protective effect of CTRP9 on the cardiac lipotoxicity. These results indicate that CTRP9 exerted anti‐myocardial lipotoxicity properties and inhibited cardiac hypertrophy probably through the LKB1/AMPK signalling pathway.     

Inflammation, gene mutation and photoimmunosuppression in response to UVR-induced oxidative damage contributes to photocarcinogenesis

Ultraviolet (UV) radiation causes inflammation, gene mutation and immunosuppression in the skin. These biological changes are responsible for photocarcinogenesis. UV radiation in sunlight is divided into two wavebands, UVB and UVA, both of which contribute to these biological changes, and therefore probably to skin cancer in humans and animal models. Oxidative damage caused by UV contributes to inflammation, gene mutation and immunosuppression. This article reviews evidence for the hypothesis that UV oxidative damage to these processes contributes to photocarcinogenesis. UVA makes a larger impact on oxidative stress in the skin than UVB by inducing reactive oxygen and nitrogen species which damage DNA, protein and lipids and which also lead to NAD+ depletion, and therefore energy loss from the cell. Lipid peroxidation induces prostaglandin production that in association with UV-induced nitric oxide production causes inflammation. Inflammation drives benign human solar keratosis (SK) to undergo malignant conversion into squamous cell carcinoma (SCC) probably because the inflammatory cells produce reactive oxygen species, thus increasing oxidative damage to DNA and the immune system. Reactive oxygen or nitrogen appears to cause the increase in mutational burden as SK progress into SCC in humans. UVA is particularly important in causing immunosuppression in both humans and mice, and UV lipid peroxidation induced prostaglandin production and UV activation of nitric oxide synthase is important mediators of this event. Other immunosuppressive events are likely to be initiated by UV oxidative stress. Antioxidants have also been shown to reduce photocarcinogenesis. While most of this evidence comes from studies in mice, there is supporting evidence in humans that UV-induced oxidative damage contributes to inflammation, gene mutation and immunosuppression. Available evidence implicates oxidative damage as an important contributor to sunlight-induced carcinogenesis in humans.     

Ovarian carcinoma cells inhibit T cell proliferation: suppression of IL-2 receptor beta and gamma expression and their JAK-STAT signaling pathway

Deficient T cell immune function and intracellular signaling in cancer patients may result from effects of tumors or their products on lymphocytes. Recently, it was demonstrated that several ovarian carcinoma cell lines could produce soluble factors that inhibited T cell proliferation. The aim of this study is to assess the effect of supernatants from 3 ovarian carcinoma cell lines (OVCAR3, CAOV3, SKOV3) on signal transduction elements that are linked to the IL-2R and its JAK-STAT pathway. A profound inhibition of proliferation, lower level of IFN-gamma and higher level of IL-10 gene expression were observed when CD8+ T cells were co-cultured with supernatants from 3 ovarian carcinoma cell lines. Cell cycle studies on inhibited CD8+ T cells showed most of them were growth arrested in G0/G1 phase. Western blot analysis showed that tumor supernatants suppressed expression of JAK3 and tyrosine phosphorylation of STAT5. JAK1 was not altered and the inhibition of STAT3 only appeared in OVCAR3 cells. Tumor supernatants also partially blocked induction of IL-2R beta and gamma chains expression. These findings suggest that ovarian carcinoma cells may suppress T cell proliferation through inhibition IL-2 dependent signaling pathways, which may be a mechanism of ovarian carcinoma induced immunosuppression.     

Differential sensitivity to apoptosis between the human small and large intestinal mucosae: linkage with segment-specific regulation of BCL-2 homologs and involvement of signaling pathways.

The small and large intestines differ in their expression profiles of Bcl-2 homologs. Intestinal segment-specific Bcl-2 homolog expression profiles are acquired as early as by mid-gestation (18-20 weeks) in man. In the present study, we examined the question whether such distinctions underlie segment-specific control mechanisms of intestinal cell survival. Using mid-gestation human jejunum and colon organotypic cultures, we analyzed the impact of growth factors (namely insulin; 10 microg/ml) and pharmacological compounds that inhibit signal transduction molecules/pathways (namely tyrosine kinases, Fak, P13-K/Akt, and MEK/Erk) on cell survival and Bcl-2 homolog expression (anti-apoptotic: Bcl-2, Bcl-X(L), Mcl-1; pro-apoptotic: Bax, Bak, Bad). The relative activation levels of p125Fak, p42Erk-2, and p57Akt were analyzed as well. Herein, we report that (1) the inhibition of signal transduction molecules/pathways revealed striking differences in their impact on cell survival in the jejunum and colon (e.g., the inhibition of p125Fak induced apoptosis with a significantly greater extent in the jejunum [approximately 43%] than in the colon [approximately 24%]); (2) sharp distinctions between the two segments were noted in the modulatory effects of the various treatments on Bcl-2 homolog steady-state levels (e.g., inhibition of tyrosine kinase activities in the jejunum down-regulated all anti-apoptotics analyzed while increasing Bax, whereas the same treatment in the colon down-regulated Bcl-X(L) only and increased all pro-apoptotics); and (3) in addition to their differential impact on cell survival and Bcl-2 homolog expression, the MEK/Erk and P13-K/Akt pathways were found to be distinctively regulated in the jejunum and colon mucosae (e.g., insulin in the jejunum increased p42Erk-2 activation without affecting that of p57Akt, whereas the same treatment in the colon decreased p42Erk-2 activation while increasing that of p57Akt). Altogether, these data show that intestinal cell survival is characterized by segment-specific susceptibilities to apoptosis, which are in turn linked with segmental distinctions in the involvement of signaling pathways and the regulation of Bcl-2 homolog steady-state levels. Therefore, these indicate that cell survival is subject to segment-specific control mechanisms along the proximal-distal axis of the intestine.     

T cell receptor stimulation, reactive oxygen species, and cell signaling

In the immune system, much of the focus on reactive oxygen species (ROS) has been regarding their role in antimicrobial defense as part of the innate immune system. In addition to this role, it is now becoming clear that ROS are used by cells of the adaptive immune system as regulators of signal transduction by cell surface receptors. The activation of T lymphocytes through their specific antigen receptor [T cell receptor (TCR)] is vital in regulating the immune response. Much experimental evidence has suggested that activation of T cells is redox dependent and recent studies have shown that engagement of the TCR induces rapid production of ROS. This review examines the evidence for TCR-stimulated generation of ROS and discusses the role(s) of receptor-stimulated ROS production in T cell signal transduction and gene expression.     

Osmotic regulation of bile acid transport, apoptosis and proliferation in rat liver

Changes in mammalian cell volume as induced by either anisoosmolarity, hormones, nutrients or oxidative stress critically contribute to the regulation of metabolism, membrane transport, gene expression and the susceptibility to cellular stress. Osmosensing, i.e. the registration of cell volume changes, triggers signal transduction pathways towards effector pathways (osmosignaling) which link alterations of cell volume to changes in cell function. This review summarizes our own work on the understanding of how osmosensing and osmosignaling integrate into the overall context of bile acid transport, growth factor signaling and the execution of apoptotic programs.     

Phosphosite-specific regulation of the oxidative-stress response of Paracoccidioides brasiliensis: a shotgun phosphoproteomic analysis

Paracoccidioides brasiliensis, a thermally dimorphic fungus, is the causative agent of paracoccidioidomycosis, a systemic mycosis that is widespread in Latin America. This fungus is a facultative intracellular pathogen able to survive and replicate inside non-activated macrophages. Therefore, the survival of P. brasiliensis inside the host depends on the ability to adapt to oxidative stress induced by immune cells, especially alveolar macrophages. For several years, reactive oxygen species (ROS) were only associated with pathological processes. Currently, a plethora of roles for ROS in cell signaling have emerged. We have previously reported that low ROS concentrations cause cell proliferation in the human pathogenic fungus P. brasiliensis. In the present report, we investigated the influence of phosphorylation events in that process. Using a mass spectrometry-based approach, we mapped 440 phosphorylation sites in 230 P. brasiliensis proteins and showed that phosphorylation at different sites determines fungal responses to oxidative stress, which are regulated by phosphatases and kinases activities. Furthermore, we present additional evidence for a functional two-component signal transduction system in P. brasiliensis. These findings will help us to understand the phosphorylation events involved in the oxidative stress response.     

Protective effect of propolis against oxidative stress and immunosuppression induced by oxytetracycline in rainbow trout (Oncorhynchus mykiss, W.)

The aim of this study was to investigate the effects of propolis on oxytetracycline (OTC)-induced oxidative stress and immunosuppression in fish. OTC (100 mg per kg?1 body weight) was orally administered to fish for 14 days. A significant elevation in the level of malondialdehyde, as an index of lipid peroxidation, and reductions in antioxidant enzyme activities (superoxide dismutase, catalase, and glutathione peroxidase) and low molecular weight antioxidant (reduced glutathione) levels were observed in the blood, liver, kidney, spleen, and heart tissues of OTC-treated fish. OTC also had a suppressive effect on specific and non-specific immune system parameters, such as leucocyte counts, oxidative radical production (nitrobluetetrazolium activity), total plasma protein and immunoglobulin levels, and phagocytic activity. Pre-treatment, post-treatment, and simultaneous treatment with propolis (50 mg per kg?1 body weight, orally) attenuated the OTC-induced oxidative stress by significantly decreasing the levels of malondialdehyde in tissues. In addition, propolis significantly increased the level of reduced glutathione and the catalase, glutathione peroxidase, and superoxide dismutase activities. Upon the administration of propolis, the suppressed immune system parameters were significantly increased in fish treated with OTC. The present results suggest that pre-treatment, post-treatment, and simultaneous administration of propolis might alleviate OTC-induced oxidative stress and immunosuppression.     

Osmosensing and signaling in the regulation of mammalian cell function

Volume changes of mammalian cells as induced by either anisoosmolarity or under isoosmotic conditions by hormones, substrates and oxidative stress critically contribute to the regulation of metabolism, gene expression and the susceptibility to stress. Osmosensing (i.e. the registration of cell volume) triggers signal transduction pathways towards effector sites (osmosignaling), which link alterations of cell volume to a functional outcome. This minireview summarizes recent progress in the understanding of how osmosensing and osmosignaling integrate into the overall context of growth factor signaling and the execution of apoptotic programs.     

Independent pathways leading to apoptotic cell death, oxidative burst and defense gene expression in response to elicitin in tobacco cell suspension culture.

We characterized pharmacologically the hypersensitive cell death of tobacco BY-2 cells that followed treatments with Escherichia coli preparations of INF1, the major secreted elicitin of the late blight pathogen Phytophthora infestans. INF1 elicitin treatments resulted in fragmentation and 180 bp laddering of tobacco DNA as early as 3 h post-treatment. INF1 elicitin also induced rapid accumulation of H2O2 typical of oxidative burst, and the expression of defense genes such as phenylalanine ammonia-lyase (PAL) gene at 1 h and 3 h after elicitin treatment, respectively. To investigate the involvement of the oxidative burst and/or the expression of defense genes in the signal transduction pathways leading to hypersensitive cell death, we analyzed the effect of several chemical inhibitors of signal transduction pathways on the various responses. The results indicated that (a) the cell death required serine proteases, Ca2+ and protein kinases, (b) the oxidative burst was involved in Ca2+ and protein kinase mediated pathways, but elicitin-induced AOS was neither necessary nor sufficient for cell death and PAL gene expression, and (c) the signaling pathway of PAL gene expression required protein kinases. These results suggest that the three signal transduction pathways leading to cell death, oxidative burst and expression of defense genes branch in the early stages that follow elicitin recognition by tobacco cells.     

Lymphocyte enzymatic antioxidant responses to oxidative stress following high-intensity interval exercise

The purposes of this study were to 1) examine the immune and oxidative stress responses following high-intensity interval training (HIIT); 2) determine changes in antioxidant enzyme gene expression and enzyme activity in lymphocytes following HIIT; and 3) assess pre-HIIT, 3-h post-HIIT, and 24-h post-HIIT lymphocyte cell viability following hydrogen peroxide exposure in vitro. Eight recreationally active males completed three identical HIIT protocols. Blood samples were obtained at preexercise, immediately postexercise, 3 h postexercise, and 24 h postexercise. Total number of circulating leukocytes, lymphocytes, and neutrophils, as well as lymphocyte antioxidant enzyme activities, gene expression, cell viability (CV), and plasma thiobarbituric acid-reactive substance (TBARS) levels, were measured. Analytes were compared using a three (day) × four (time) ANOVA with repeated measures on both day and time. The a priori significance level for all analyses was P < 0.05. Significant increases in superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX) activities were observed in lymphocytes following HIIT. No significant increases in lymphocyte SOD, CAT, or GPX gene expression were found. A significant increase in TBARS was found immediately post-HIIT on days 1 and 2. Lymphocyte CV in vitro significantly increased on days 2 and 3 compared with day 1. Additionally, there was a significant decrease in CV at 3 h compared with pre- and 24 h postexercise. These findings indicate lymphocytes respond to oxidative stress by increasing antioxidant enzyme activity. Additionally, HIIT causes oxidative stress but did not induce a significant postexercise lymphocytopenia. Analyses in vitro suggest that lymphocytes may become more resistant to subsequent episodes of oxidative stress. Furthermore, the analysis in vitro confirms that lymphocytes are more vulnerable to cytotoxic molecules during recovery from exercise.     

MAPK/JNK signalling: a potential autophagy regulation pathway

Autophagy refers to a lysosomal degradative pathway or a process of self-cannibalization. This pathway maintains nutrients levels for vital cellular functions during periods of starvation and it provides cells with survival advantages under various stress situations. However, the mechanisms responsible for the induction and regulation of autophagy are poorly understood. The c-Jun NH2-terminal kinase (JNK) signal transduction pathway functions to induce defence mechanisms that protect organisms against acute oxidative and xenobiotic insults. This pathway has also been repeatedly linked to the molecular events involved in autophagy regulation. The present review will focus on recent advances in understanding of the relationship between mitogen-activated protein kinase (MAPK)/JNK signalling and autophagic cell death.