Upregulation Of Stress Response Genes Research Articles

The Juvenile Phase of Maize Sees Upregulation of Stress-Response Genes and Is Extended by Exogenous Jasmonic Acid.

As maize (Zea mays) plants undergo vegetative phase change from juvenile to adult, they both exhibit heteroblasty, an abrupt change in patterns of leaf morphogenesis, and gain the ability to produce flowers. Both processes are under the control of microRNA156 (miR156), whose levels decline at the end of the juvenile phase. Gain of the ability to flower is conferred by the expression of miR156 targets that encode SQUAMOSA PROMOTER-BINDING transcription factors, which, when derepressed in the adult phase, induce the expression of MADS box transcription factors that promote maturation and flowering. How gene expression, including targets of those microRNAs, differs between the two phases remains an open question. Here, we compare transcript levels in primordia that will develop into juvenile or adult leaves to identify genes that define these two developmental states and may influence vegetative phase change. In comparisons among successive leaves at the same developmental stage, plastochron 6, three-fourths of approximately 1,100 differentially expressed genes were more highly expressed in primordia of juvenile leaves. This juvenile set was enriched in photosynthetic genes, particularly those associated with cyclic electron flow at photosystem I, and in genes involved in oxidative stress and retrograde redox signaling. Pathogen- and herbivory-responsive pathways including salicylic acid and jasmonic acid also were up-regulated in juvenile primordia; indeed, exogenous application of jasmonic acid delayed both the appearance of adult traits and the decline in the expression of miR156-encoding loci in maize seedlings. We hypothesize that the stresses associated with germination promote juvenile patterns of differentiation in maize.

Compression Induced Stress Response of Nucleus Pulposus Cells in 3D Collagen Gel

Introduction The cause of degenerative disc disease is multi-factorial and is often associated with mechanical loading. However, mechanism of the cells' response to compressive loading is not fully known. It was demonstrated that in mechanical force can induce chondrocytes to upregulate expression of heat shock proteins (HSPs) as a form of cellular stress response.1 Similarly, stretching annulus fibrosus cells was found to upregulate endoplasmic reticulum stress markers.2 However, it is unclear that whether cells in the intervertebral disc (IVD) would experience compressive force as cellular stress in 3D environment. The objective of this study is to investigate the stress response of nucleus pulposus cells (NPCs) during compression in 3D culture environment. Materials and Methods To simulate the condition in the IVD, NPCs were isolated from bovine caudal disc and encapsulated in 3D collagen gel as described previously.3 The culture system was shown to be able to maintain NPCs' phenotype.4 The cell-encapsulated gels were cultured for 4 days and then compressed with different types of loading (static and dynamic loading), different strains (10% - 70%) and different durations (2h - 8h). Samples were retrieved to study the expressions of stress response genes in two groups: Heat Shock Response (HSP70, HSP27, HSP90 and HSF1) and Unfolded Protein Response (GRP78, GRP94, ATF4 and CHOP). Results Encapsulation in 3D collagen gel was found to upregulate HSP70 in NPCs. Compression further induced stress response of NPCs which depended on multiple factors. The upregulation of stress response genes was generally insignificant with increasing loading strain in short duration. However the upregulations increased with longer duration where increasing loading duration has a larger effect in the upregulation of HSP70 and HSP27. When high static strain was applied, the expressions of Heat Shock Response genes were continued to be upregulated with time after the load was removed while Unfolded Protein Response genes were downregulated with incubation duration. The changes in expression with incubation duration show that cellular stress response may play a role in NPC survival and protein homeostasis post-loading. Conclusion The NPCs demonstrated cellular stress response during compression in collagen gel. The expression of stress response genes were mainly affected by loading duration and incubation duration after loading. This study helps us to understand how the NPCs cope with mechanical stress. Acknowledgments This project was funded by TBRS (T12–708/12-N). References Kaarniranta K, Elo M, Sironen R, et al. Hsp70 accumulation in chondrocytic cells exposed to high continuous hydrostatic pressure coincides with mRNA stabilization rather than transcriptional activation. Proc Natl Acad Sci U S A 1998;95(5):2319–2324 Zhang YH, Zhao CQ, Jiang LS, Dai LY. Cyclic stretch-induced apoptosis in rat annulus fibrosus cells is mediated in part by endoplasmic reticulum stress through nitric oxide production. Eur Spine J 2011;20(8):1233–1243 Chan BP, Hui TY, Yeung CW, Li J, Mo I, Chan GC. Self-assembled collagen-human mesenchymal stem cell microspheres for regenerative medicine. Biomaterials 2007;28(31):4652–4666 Yuan M, Leong KW, Chan BP. Three-dimensional culture of rabbit nucleus pulposus cells in collagen microspheres. Spine J 2011;11(10):947–960

Identification and expression of proteolysis-response genes forMacrophthalmus japonicusexposure to irgarol toxicity

Macrophthalmus japonicus is one of the most abundant macrobenthic animals in estuarine zone, which are transition areas between freshwater and marine environments. The crabs are potential bio-indicators reflecting aquatic sediment toxicity. So far, limited genetic research has focused on this group of animals largely due to the limited genomic information. In this study, we performed de novo transcriptome sequencing to produce the most comprehensive expressed sequence tag resource for M. japonicus , and identified stress response genes in M. japonicus exposed to antifouling biocides irgarol. Using 454 pyrosequencing, a total of 887 690 reads were obtained, which were assembled into 24 217 high-quality expressed sequence tags. These contigs were then clustered into 17 289 isotigs and further grouped into 12 923 isogroups. About 48% of the isogroups showed significant matches to known proteins based on sequence similarity. Moreover, irgarol toxicity induced up-regulation of stress response genes, associated with proteolysis and oxidation–reduction in biological process by Gene ontology analyses. Our data provide the most comprehensive transcriptome resource available for M. japonicus . This resource allowed us to identify genes associated with proteolysis processes, which facilitated the quantitative analysis of differential gene expression respond to irgarol toxicity. These data would provide foundation for future genetic and genomic studies of crustacean species and monitoring to freshwater as well as marine ecosystems.

TraeALDH7B1-5A, encoding aldehyde dehydrogenase 7 in wheat, confers improved drought tolerance in Arabidopsis.

TraeALDH7B1 - 5A , encoding aldehyde dehydrogenase 7 in wheat, conferred significant drought tolerance to Arabidopsis , supported by molecular biological and physiological experiments. Drought stress significantly affects wheat yields. Aldehyde dehydrogenase (ALDH) is a family of enzymes catalyzing the irreversible conversion of aldehydes into acids to decrease the damage caused by abiotic stresses. However, no wheat ALDH member has been functionally characterized to date. Here, we obtained a differentially expressed EST encoding ALDH7 from a cDNA-AFLP library of wheat that was treated with polyethylene glycol 6000. The three full-length homologs of TraeALDH7B1 were isolated by searching the NCBI database and by homolog-based cloning method. Using nulli-tetrasomic lines we located them on wheat chromosomes 5A, 5B and 5D, and named them as TraeALDH7B1-5A, -5B and -5D, respectively. Gene expression profiles indicated that the expressions of all three genes were induced in roots, leaves, culms and spikelets under drought and salt stresses. Enzymatic activity analysis showed that TraeALDH7B1-5A had acetaldehyde dehydrogenase activity. For further functional analysis, we developed transgenic Arabidopsis lines overexpressing TraeALDH7B1-5A driven by the cauliflower mosaic virus 35S promoter. Compared with wild type Arabidopsis, 35S::TraeALDH7B1-5A plants significantly enhanced the tolerance to drought stress, which was demonstrated by up-regulation of stress responsive genes and physiological evidence of primary root length, maintenance of water retention and contents of chlorophyll and MDA. The combined results indicated that TraeALDH7B1-5A is an important drought responsive gene for genetic transformation to improve drought tolerance in crops.

Pea lectin receptor-like kinase functions in salinity adaptation without yield penalty, by alleviating osmotic and ionic stresses and upregulating stress-responsive genes.

Lectin receptor-like kinases (LecRLKs) are members of RLK family composed of lectin-like extracellular recognition domain, transmembrane domain and cytoplasmic kinase domain. LecRLKs are plasma membrane proteins believed to be involved in signal transduction. However, most of the members of the protein family even in plants have not been functionally well characterized. Herein, we show that Pisum sativum LecRLK (PsLecRLK) localized in plasma membrane systems and/or other regions of the cell and its transcript upregulated under salinity stress. Overexpression of PsLecRLK in transgenic tobacco plants confers salinity stress tolerance by alleviating both the ionic as well the osmotic component of salinity stress. The transgenic plants show better tissue compartmentalization of Na(+) and higher ROS scavenging activity which probably results in lower membrane damage, improved growth and yield maintenance even under salinity stress. Also, expression of several genes involved in cellular homeostasis is perturbed by PsLecRLK overexpression. Alleviation of osmotic and ionic components of salinity stress along with reduced oxidative damage and upregulation of stress-responsive genes in transgenic plants under salinity stress conditions could be possible mechanism facilitating enhanced stress tolerance. This study presents PsLecRLK as a promising candidate for crop improvement and also opens up new avenue to investigate its signalling pathway.

Arabidopsis flower specific defense gene expression patterns affect resistance to pathogens.

We investigated whether the Arabidopsis flower evolved protective measures to increase reproductive success. Firstly, analyses of available transcriptome data show that the most highly expressed transcripts in the closed sepal (stage 12) are enriched in genes with roles in responses to chemical stimuli and cellular metabolic processes. At stage 15, there is enrichment in transcripts with a role in responses to biotic stimuli. Comparative analyses between the sepal and petal in the open flower mark an over-representation of transcripts with a role in responses to stress and catalytic activity. Secondly, the content of the biotic defense-associated phytohormone salicylic acid (SA) in sepals and petals is significantly higher than in leaves. To understand whether the high levels of stress responsive transcripts and the higher SA content affect defense, wild-type plants (Col-0) and transgenic plants defective in SA accumulation (nahG) were challenged with the biotrophic fungus Golovinomyces cichoracearum, the causal agent of powdery mildew, and the necrotrophic fungus Botrytis cinerea. NahG leaves were more sensitive than those of Col-0, suggesting that in leaves SA has a role in the defense against biotrophs. In contrast, sepals and petals of both genotypes were resistant to G. cichoracearum, indicating that in the flower, resistance to the biotrophic pathogen is not critically dependent on SA, but likely dependent on the up-regulation of stress-responsive genes. Since sepals and petals of both genotypes are equally susceptible to B. cinerea, we conclude that neither stress-response genes nor increased SA accumulation offers protection against the necrotrophic pathogen. These results are interpreted in the light of the distinctive role of the flower and we propose that in the early stages, the sepal may act as a chemical defense barrier of the developing reproductive structures against biotrophic pathogens.

OsBAT1 Augments Salinity Stress Tolerance by Enhancing Detoxification of ROS and Expression of Stress-Responsive Genes in Transgenic Rice

Coping with salinity-induced reduction in crop yield is essential for food security. Human leukocyte antigen-B associated transcript 1 (BAT1) also called as UAP56 is a DExD/H-box protein involved in messenger RNA (mRNA) splicing. Function of plant homologue of BAT1, especially its involvement in stress tolerance, has not been reported so far. Here, we demonstrate the localization of rice BAT1 (OsBAT1) in the nucleus and in the plasma membrane and its novel function in salinity stress tolerance in rice (Oryza sativa L. cv. IR64). Rice overexpressing OsBAT1 (T1 and T2 generations) show tolerance to high salinity (200 mM NaCl) stress. The T1 transgenics exhibited higher levels of biochemical parameters such as water and chlorophyll contents, net photosynthetic rate, stomatal conductance and intercellular CO2 content as compared to null-segregant (control) plants. The activities of ascorbate peroxidase, guaiacol peroxidase, malondialdehyde and glutathione reductase were significantly higher in transgenics indicating the presence of an efficient antioxidant defence system which helps to cope with salinity-induced oxidative damages. Agronomic parameters were also higher in transgenics as compared to control. Microarray analysis of OsBAT1 overexpressing transgenic lines revealed up-regulation of stress-responsive genes of different pathways including the spliceosome. Our results provide the first direct evidence for a promising function of OsBAT1 in mediating salinity stress response/tolerance in rice.

Alleviation of drought stress in mung bean by strain Pseudomonas aeruginosa GGRJ21

Mung bean (Vigna radiata (L.) R. Wilczek), a widely cultivated pulse crops in India, experiences severe drought stress during the cultivation period. Apart from the conventional plant breeding and transgenic approaches, the application of plant growth-promoting rhizobacteria (PGPR) has always been a promising approach to improve abiotic stress tolerance in crop plants. The aim of the present study was to investigate the role of mung bean rhizosphere-associated Pseudomonas aeruginosa GGRJ21 strain on drought stress alleviation in the host plant. Fluorescent pseudomonads were isolated from mung bean rhizosphere by employing a culture-dependent approach. The role of osmotic stress tolerant P. aeruginosa GGRJ21 on drought stress alleviation in host plants was further examined in both the green house and field conditions. An elevated production of reactive oxygen species scavenging enzymes and cellular osmolytes; increased root length, shoot length, dry weight, relative water content; and a stronger upregulation of three drought stress-responsive genes, i.e., dehydration-responsive element binding protein (DREB2A), catalase (CAT1), and dehydrin (DHN) were observed in GGRJ21 inoculated plants in comparison with the uninoculated control plants tested under drought conditions. The field experimental data show an increase in biomass and better growth and development in inoculated and stressed plants when compared with untreated and stressed plants. P. aeruginosa GGRJ21 strain was found to elicit water stress tolerance in mung bean plants by accelerating the accumulation of inherent levels of antioxidant enzymes, cell osmolytes, and consistently expediting the upregulation of stress responsive genes in PGPR-treated plants under water stress conditions.

PECA: a novel statistical tool for deconvoluting time-dependent gene expression regulation.

Protein expression varies as a result of intricate regulation of synthesis and degradation of messenger RNAs (mRNA) and proteins. Studies of dynamic regulation typically rely on time-course data sets of mRNA and protein expression, yet there are no statistical methods that integrate these multiomics data and deconvolute individual regulatory processes of gene expression control underlying the observed concentration changes. To address this challenge, we developed Protein Expression Control Analysis (PECA), a method to quantitatively dissect protein expression variation into the contributions of mRNA synthesis/degradation and protein synthesis/degradation, termed RNA-level and protein-level regulation respectively. PECA computes the rate ratios of synthesis versus degradation as the statistical summary of expression control during a given time interval at each molecular level and computes the probability that the rate ratio changed between adjacent time intervals, indicating regulation change at the time point. Along with the associated false-discovery rates, PECA gives the complete description of dynamic expression control, that is, which proteins were up- or down-regulated at each molecular level and each time point. Using PECA, we analyzed two yeast data sets monitoring the cellular response to hyperosmotic and oxidative stress. The rate ratio profiles reported by PECA highlighted a large magnitude of RNA-level up-regulation of stress response genes in the early response and concordant protein-level regulation with time delay. However, the contributions of RNA- and protein-level regulation and their temporal patterns were different between the two data sets. We also observed several cases where protein-level regulation counterbalanced transcriptomic changes in the early stress response to maintain the stability of protein concentrations, suggesting that proteostasis is a proteome-wide phenomenon mediated by post-transcriptional regulation.

Global transcriptome analysis of Escherichia coli exposed to immobilized anthraquinone-2-sulfonate and azo dye under anaerobic conditions

The immobilization of quinone compounds is regarded as a promising strategy to accelerate anaerobic decolorization of xenobiotic compounds azo dyes in the presence of quinone-reducing microorganisms. However, little is known about the basic response of these microorganisms to immobilized quinones in the presence of azo dyes. In the present study, whole-genome DNA microarrays were used to investigate a quinone-reducing bacterium Escherichia coli K-12 transcription response to immobilized anthraquinone-2-sulfonate (AQSim) reduction and azo dye acid red 18 (AR 18) decolorization. Transcriptome analysis showed that AQSim was more accessible for the cells of E. coli K-12 than AR 18. Despite there being some differences between AQSim and soluble AQS mediated decolorization of AR 18, AQSim reduction and AR 18 decolorization, more similarity could be observed in the four processes. Among over 60 % shared genes, several groups of genes exhibited high expression levels, including those genes encoding terminal reductases, menaquinone biosynthesis, formate dehydrogenases and outer membrane proteins. Especially, nrfABCD, frdBCD and dsmABC encoding terminal reductases were significantly upregulated. Further gene deletion experiments demonstrated that the above three groups of genes were involved in AQSim-mediated AR 18 decolorization. In addition, significant upregulation of stress response genes was observed, which indicated the adaptation of E. coli K-12 to AQSim and AR 18 exposures.

Inositol Pyrophosphates Regulate Cell Growth and the Environmental Stress Response by Activating the HDAC Rpd3L

Cells respond to stress and starvation by adjusting their growth rate and enacting stress defense programs. In eukaryotes this involves inactivation of TORC1, which in turn triggers downregulation of ribosome and protein synthesis genes and upregulation of stress response genes. Here we report that the highly conserved inositol pyrophosphate (PP-IP) second messengers (including 1-PP-IP5, 5-PP-IP4, and 5-PP-IP5) are also critical regulators of cell growth and the general stress response, acting in parallel with the TORC1 pathway to control the activity of the class I histone deacetylase Rpd3L. In fact, yeast cells that cannot synthesize any of the PP-IPs mount little to no transcriptional response to osmotic, heat, or oxidative stress. Furthermore, PP-IP-dependent regulation of Rpd3L occurs independently of the role individual PP-IPs (such as 5-PP-IP5) play in activating specialized stress/starvation response pathways. Thus, the PP-IP second messengers simultaneously activate and tune the global response to stress and starvation signals.

Effects of postharvest treatments on gene expression in Prunus persica fruit: Normal and altered ripening

Peach (Prunus persica) fruit have a short shelf-life, and the most common method employed to delay ripening and increase their postharvest life is cold storage. However, after extended storage at low temperature some cultivars have alterated ripening processes, resulting in a lack of juice and a woolly texture. To improve our understanding of the molecular mechanisms involved in the responses of peach fruit to cold storage we determined gene expression changes of fruit (cv. O’Henry) under different postharvest conditions: ripening (5days at 21°C), cold storage (21days at 4°C) and induction of woolliness (21days at 4°C followed by 5days at 21°C).Cluster analyses of genes differentially expressed between treatments revealed unique patterns associated with biological processes that operate during postharvest treatments. Genes up-regulated during postharvest ripening and woolliness include components of ethylene, and aroma biosynthesis as well as oxidative stress response. During cold storage treatment and woolliness, several genes linked to the oxidative stress response increased in abundance, suggesting changes in redox status. Quantitative RT-PCR analysis showed a sequential increase levels of mRNAs encoding key components of cellular stress response. Moreover, after 21days of cold storage, expression of genes encoding oxidoreductase, catalase, superoxide dismutase and gluthatione reductase was still significantly higher than before cold treatment, suggesting that fruit cells were able to respond to the increased production of ROS that was induced by extended cold storage. In the woolly fruit, up-regulation of stress response genes was accompanied by down-regulation of key components of metabolic pathways that are active during peach ripening. The altered expression pattern of these genes might account for the abnormal ripening of woolly fruit.

Imatinib: the controversial discussion on cardiotoxicity induced by endoplasmic reticulum (ER) stress

The safety profile of imatinib, and especially its relevance to cardiotoxicity, has been controversially discussed in recent years (Kerkela et al. 2006; Herman et al. 2011; Wolf et al. 2010; Ribeiro et al. 2008). Therefore, the editors are pleased that Eirini Thanopoulou and Ian Judson from the Royal Marsden NHS foundation Trust in London contributed a comprehensive review on the long-term tolerability of imatinib (Gleevec) (Thanopoulou and Judson 2012). The introduction of imatinib represents one of the most prominent success stories in cancer therapy of the past decade. A reciprocal translocation creating the Philadelphia chromosome is involved in pathogenesis of more than 90% of the cases of chronic myelogenous leukaemia (CML) (Van Etten 2004). This translocation creates the Bcr-Abl fusion protein of the nonreceptor tyrosine kinase c-Abl and the multidomain protein Bcr, leading to a constitutively active cytosolic protein that activates several signalling pathways involved in proliferation and antiapoptosis. Imatinib inhibits the fusion protein, Bcr-Abl resulting in remission in a high fraction of patients with CML. In addition, imatinib is efficient in gastrointestinal stromal tumour (GIST), because—besides its effect on Bcr-Abl—it also inhibits KIT proteins that play a role in progression of GIST and other tumours (Joensuu et al. 2001; Micke et al. 2003, 2004). The success stories of imatinib, and the Her-2-targeting antibody, trastuzumab, led to the novel concept of identifying and targeting the mechanisms responsible of tumour progression, in addition to identifying subtypes of carcinomas with the perspective of a biologically justified therapy (Schmidt et al. 2008, 2010; Hellwig et al. 2010; Cadenas et al. 2010; Lee et al. 2010; Brase et al. 2010; Petry et al. 2010). Previously, Kerkela et al. (2006) reported on the cardiotoxic side effect of imatinib. Individuals without any prior history of heart disease developed heart failure after treatment with imatinib (Kerkela et al. 2006). Interestingly, imatinib also showed toxic effects on cultivated cardiomyocytes, most likely due to an induction of endoplasmic reticulum (ER) stress, leading to decreased translation and a block in protein synthesis (Kerkela et al. 2006). A simultaneous upregulation of stress response genes suggested that the ER stress response is an adequate mechanism to compensate short-term cell stress. However, prolonged activation may induce cell death by pathways, including Jun N-terminal kinases (JNKs), a mechanism shown for imatinib in cardiomyocytes (Kerkela et al. 2006). Since cardiotoxicity is one of the leading topics in toxicology addressed in our journal (Sumi et al. 2011; Vavrova et al. 2011; Lee et al. 2010), we welcome the current review by Thanopoulou and Judson (2012). Overall, imatinib shows an excellent long-term tolerability and the clinical relevance of imatinib-induced cardiotoxicity may have been overestimated. The comprehensive review of Thanopoulou and Judson (2012) comes highly recommended for anyone interested in the safety profile of imatinib.

Zinc pyrithione impairs zinc homeostasis and upregulates stress response gene expression in reconstructed human epidermis

Zinc ion homeostasis plays an important role in human cutaneous biology where it is involved in epidermal differentiation and barrier function, inflammatory and antimicrobial regulation, and wound healing. Zinc-based compounds designed for topical delivery therefore represent an important class of cutaneous therapeutics. Zinc pyrithione (ZnPT) is an FDA-approved microbicidal agent used worldwide in over-the-counter topical antimicrobials, and has also been examined as an investigational therapeutic targeting psoriasis and UVB-induced epidermal hyperplasia. Recently, we have demonstrated that cultured primary human skin keratinocytes display an exquisite sensitivity to nanomolar ZnPT concentrations causing induction of heat shock response gene expression and poly(ADP-ribose) polymerase (PARP)-dependent cell death (Cell Stress Chaperones 15:309-322, 2010). Here we demonstrate that ZnPT causes rapid accumulation of intracellular zinc in primary keratinocytes as observed by quantitative fluorescence microscopy and inductively coupled plasma mass spectrometry (ICP-MS), and that PARP activation, energy crisis, and genomic impairment are all antagonized by zinc chelation. In epidermal reconstructs (EpiDerm™) exposed to topical ZnPT (0.1-2% in Vanicream™), ICP-MS demonstrated rapid zinc accumulation, and expression array analysis demonstrated upregulation of stress response genes encoding metallothionein-2A (MT2A), heat shock proteins (HSPA6, HSPA1A, HSPB5, HSPA1L, DNAJA1, HSPH1, HSPD1, HSPE1), antioxidants (SOD2, GSTM3, HMOX1), and the cell cycle inhibitor p21 (CDKN1A). IHC analysis of ZnPT-treated EpiDerm™ confirmed upregulation of Hsp70 and TUNEL-positivity. Taken together our data demonstrate that ZnPT impairs zinc ion homeostasis and upregulates stress response gene expression in primary keratinocytes and reconstructed human epidermis, activities that may underlie therapeutic and toxicological effects of this topical drug.

Matrix metalloproteinases are possible mediators for the development of alimentary tract mucositis in the dark agouti rat

Alimentary tract (AT) mucositis is a serious and debilitating side-effect of cancer therapy primarily characterized by damage of the mucous membranes throughout the AT. It is well established that this damage is a result of up-regulation of stress response genes and pro-inflammatory cytokines. Matrix metalloproteinases (MMPs) have been shown to function in several of the pathways known to be up-regulated in mucositis and play a key role in tissue injury and inflammation in many gastrointestinal disorders. This study aims to characterize the expression of multiple MMPs including MMP-1, -2, -3, -9 and -12 and their inhibitors, tissue inhibitor of metalloproteinase (TIMP)-1 and -2, in a rat model of irinotecan-induced mucositis. Dark agouti rats were administered a single 200 mg/kg intraperitoneal dose of irinotecan and killed at 1, 6, 24, 48, 72, 96 and 144 h following treatment. Hematoxylin and eosin staining, immunohistochemistry and realtime polymerase chain reaction were used to assess histopathological damage and MMP expression in the jejunum and colon. Marked histopathological evidence of mucositis was observed in the jejunum and colon as early as six hours following irinotecan treatment. A significant alteration in both gene expression and tissue levels of MMPs and TIMPs was noted following irinotecan. The increase in MMP-2, -3, -9 and -12 levels was associated with inflammatory infiltrate and maximum tissue damage. In contrast, MMP-1 expression correlated with tissue restitution. TIMP-1 and -2 levels were significantly altered in the jejunum following irinotecan. The augmentation in the expression profiles of MMPs and their inhibitors correlated with histopathological alterations observed in the tissue following irinotecan. This prompts the consideration of MMPs as possible mediators of chemotherapy-induced mucositis.

Bystander Cell Death and Stress Response is Inhibited by the Radical Scavenger α1-Microglobulin in Irradiated Cell Cultures

Alpha-particle irradiation of cells damages not only the irradiated cells but also nontargeted bystander cells. It has been proposed that the bystander effect is caused by oxidants and free radicals generated by the radiation. Recent studies have shown that α(1)-microglobulin protects against cell damage caused by oxidants and free radicals. Using a novel experimental system that allows irradiation of 0.02% of a human hepatoma monolayer, leaving 99.98% as bystander cells, we investigated the influence of oxidative stress and the cell-protective effects of α(1)-microglobulin during α-particle irradiation. The results showed an increase in cell death in both irradiated cells and bystander cells. A significant increase in apoptosis, oxidation markers and expression of the stress response genes heme oxygenase 1, superoxide dismutase, catalase, glutathione peroxidase 1, p21 and p53 were observed. Addition of α(1)-microglobulin reduced the amount of dead cells and inhibited apoptosis, formation of oxidation markers, and up-regulation of stress response genes. The results emphasize the role of oxidative stress in promoting bystander effects. Furthermore, the results suggest that α(1)-microglobulin protects nonirradiated cells by eliminating oxidants and free radicals generated by radiation and imply that α(1)-microglobulin can be used in radiation therapy of tumors to minimize damage to surrounding tissues.

Matrix metalloproteinases: key regulators in the pathogenesis of chemotherapy-induced mucositis?

Chemotherapy is an effective anticancer treatment; however, it induces mucositis in a wide range of patients. Mucositis is the term used to describe the damage caused by radiation and chemotherapy to mucous membranes of the alimentary tract. This damage causes pain and ulceration, vomiting, bloating and diarrhoea, depending on the area of the alimentary tract affected. Although treatment is available for a small subset of patients suffering from mucositis, the majority rely on pain relief as their only treatment option. Much progress has been made in recent years into understanding the pathobiology underlying the development of mucositis. It is well established that chemotherapy causes prominent small intestinal and colonic damage as a result of up-regulation of stress response genes and pro-inflammatory cytokines. However, better understanding of the mediators of this damage is still required in order to target appropriate treatment strategies. Possible mediators of mucositis which have not been well researched are the matrix metalloproteinases (MMPs). MMPs have been shown to function in several of the pathways which are known to be up-regulated in mucositis and contribute to tissue injury and inflammation in many pathological conditions. This prompts the consideration of MMPs as possibly being key mediators in mucositis development.

Does an Extract of Carob (Ceratonia siliqua L.) Have Chemopreventive Potential Related To Oxidative Stress and Drug Metabolism in Human Colon Cells?

Phenolic ingredients of an aqueous carob extract are well characterized and consist of mainly gallic acid (GA). In order to assess possible chemopreventive mechanisms of carob, which can be used as a cacao substitute, effects on expression of genes related to stress response and drug metabolism were studied using human colon cell lines of different transformation state (LT97 and HT29). Stress-related genes, namely catalase (CAT) and superoxide dismutase (SOD2), were induced by carob extract and GA in LT97 adenoma, but not in HT29 carcinoma cells. Although corresponding protein products and enzyme activities were not elevated, pretreatment with carob extract and GA for 24 h reduced DNA damage in cells challenged with hydrogen peroxide (H(2)O(2)). In conclusion, carob extract and its major phenolic ingredient GA modulate gene expression and protect colon adenoma cells from genotoxic impact of H(2)O(2). Upregulation of stress-response genes could not be related to functional consequences.

Copper chelator ATN-224 inhibits endothelial function by multiple mechanisms

Copper is required for the proliferation of endothelial cells and copper-lowering therapy reduces tumour growth in animal models. It has been reported that ATN-224, a novel copper chelator, potently inhibits the activity of the copper-dependent enzyme superoxide dismutase 1 (SOD1) in endothelial cells. We performed microarray analysis of gene expression in endothelial cells exposed to ATN-224 which revealed upregulation of stress response genes including heme-oxygenase 1 (HO-1) and differential regulation of several genes previously implicated in angiogenesis including CXCR4, ANGP2, PGES2, RHAMM, ITB4 and AQP1 ( p < 0.01). These changes were confirmed on qPCR. Treatment of HUVEC with ATN-224 caused increased superoxide levels, phospho-ERK signalling, nuclear NRF1 expression, HO-1 expression and induction of the anti-apoptotic proteins P21, BCL2 and BCLXL. There was also nuclear translocation of SOD1. SOD1 RNA interference replicated the effects of ATN-224 on endothelial cell function but did not cause upregulation of HO-1 or PGES2, suggesting additional mechanisms of action of ATN-224. Downregulation of AQP1, which has been shown to have a role in angiogenesis, was seen with both ATN-224 and SOD1 siRNA. AQP1 expression could be rescued after ATN-224 by added copper. RNA interference to AQP1 inhibited endothelial proliferation and migration, confirming the role of AQP1 in endothelial cell function. Therefore regulation of AQP1 may represent an important action of copper chelation therapy.

Stress‐induced expression of an activated form of AtbZIP17 provides protection from salt stress in Arabidopsis

Membrane-associated basic-leucine zipper (bZIP) transcription factors that reside in the endoplasmic reticulum represent a newly described class of plant stress sensor/transducers. The bZIP factors are anchored in endoplasmic reticulum (ER) membranes with their C-terminal tails facing the ER lumen and their N terminii, which contain transcriptional components, facing the cytosol. In response to stress, cytosolic components of the transcription factors are released by proteolysis and move to the nucleus where they promote the up-regulation of stress response genes. One such stress sensor/transducer in Arabidopsis is AtbZIP17, which is activated in response to salt stress. With the aim of enhancing salt tolerance, a constitutively activated form of AtbZIP17, truncated before its membrane-anchor domain, was introduced into transgenic plants. When placed under the control of a constitutive promoter, the activated form of AtbZIP17 up-regulated stress response genes under unstressed conditions, but caused a substantial delay in plant development. When the activated form of AtbZIP17 was placed under the control of stress-inducible promoter, development was normal under unstressed conditions. Under salt stress conditions, the stress-inducible expression of the activated AtbZIP17 enhanced salt tolerance as demonstrated by chlorophyll bleaching and seedling survival assays.