Regulation of interleukin-8 expression in melanoma-stimulated neutrophil inflammatory response

Inflammation facilitates tumor progression including metastasis. Interleukin-8 (IL-8) is a chemokine that regulates polymorphonuclear neutrophil (PMN) mobilization and activity and we hypothesize that this cytokine influences tumor behavior. We have demonstrated that IL-8 is crucial for PMN-mediated melanoma extravasation under flow conditions. In addition, IL-8 is up-regulated in PMNs upon co-culturing with melanoma cells. Melanoma cells induce IkappaB-alpha degradation in PMNs indicating that NF-kappaB signaling is active in PMNs. Furthermore, the production of IL-8 in PMNs is NF-kappaB dependent. We have further identified that interleukin-6 (IL-6) and interleukin-1beta (IL-1beta) from PMN-melanoma co-cultures synergistically contribute to IkappaB-alpha degradation and IL-8 synthesis in PMNs. Taken together, these findings show that melanoma cells induce PMNs to secrete IL-8 through activation of NF-kappaB and suggest a model in which this interaction promotes a microenvironment that is favorable for metastasis.     

Essential functions of Alk3 during AV cushion morphogenesis in mouse embryonic hearts

Accumulated evidence has suggested that BMP pathways play critical roles during mammalian cardiogenesis and impairment of BMP signaling may contribute to human congenital heart diseases (CHDs), which are the leading cause of infant morbidity and mortality. Alk3 encodes a BMP specific type I receptor expressed in mouse embryonic hearts. To reveal functions of Alk3 during atrioventricular (AV) cushion morphogenesis and to overcome the early lethality of Alk3(-/-) embryos, we applied a Cre/loxp approach to specifically inactivate Alk3 in the endothelium/endocardium. Our studies showed that endocardial depletion of Alk3 severely impairs epithelium-mesenchymal-transformation (EMT) in the atrioventricular canal (AVC) region; the number of mesenchymal cells formed in Tie1-Cre;Alk3(loxp/loxp) embryos was reduced to only approximately 20% of the normal level from both in vivo section studies and in vitro explant assays. We showed, for the first time, that in addition to its functions on mesenchyme formation, Alk3 is also required for the normal growth/survival of AV cushion mesenchymal cells. Functions of Alk3 are accomplished through regulating expression/activation/subcellular localization of multiple downstream genes including Smads and cell-cycle regulators. Taken together, our study supports the notion that Alk3-mediated BMP signaling in AV endocardial/mesenchymal cells plays a central role during cushion morphogenesis.     

Mutations in DIIS5 and the DIIS4-S5 linker of Drosophila melanogaster sodium channel define binding domains for pyrethroids and DDT

Mutations in the DIIS4-S5 linker and DIIS5 have identified hotspots of pyrethroid and DDT interaction with the Drosophila para sodium channel. Wild-type and mutant channels were expressed in Xenopus oocytes and subjected to voltage-clamp analysis. Substitutions L914I, M918T, L925I, T929I and C933A decreased deltamethrin potency, M918T, L925I and T929I decreased permethrin potency and T929I, L925I and I936V decreased fenfluthrin potency. DDT potency was unaffected by M918T, but abolished by T929I and reduced by L925I, L932F and I936V, suggesting that DIIS5 contains at least part of the DDT binding domain. The data support a computer model of pyrethroid and DDT binding.     

A critical role for IkappaB kinase beta in metallothionein-1 expression and protection against arsenic toxicity

Arsenic is a widespread environmental toxic agent that has been shown to cause diverse tissue and cell damage and at the same time to be an effective anti-cancer therapeutic agent. The objective of this study is to explore the signaling mechanisms involved in arsenic toxicity. We show that the IkappaB kinase beta (IKKbeta) plays a crucial role in protecting cells from arsenic toxicity. Ikkbeta(-)(/)(-) mouse 3T3 fibroblasts have decreased expression of antioxidant genes, such as metallothionein 1 (Mt1). In contrast to wild type and IKKbeta-reconstituted Ikkbeta(-)(/)(-) cells, IKKbeta-null cells display a marked increase in arsenic-induced reactive oxygen species (ROS) accumulation, which leads to activation of the MKK4-c-Jun NH(2)-terminal kinase (JNK) pathway, c-Jun phosphorylation, and apoptosis. Pretreatment with the antioxidant N-acetylcysteine (NAC) and expression of MT1 in the Ikkbeta(-)(/)(-) cells prevented JNK activation; moreover, NAC pretreatment, MT1 expression, MKK4 ablation, and JNK inhibition all protected cells from death induced by arsenic. Our data show that two signaling pathways appear to be important for modulating arsenic toxicity. First, the IKK-NF-kappaB pathway is crucial for maintaining cellular metallothionein-1 levels to counteract ROS accumulation, and second, when this pathway fails, excessive ROS leads to activation of the MKK4-JNK pathway, resulting in apoptosis.     

A protocol for differential display of mRNA expression using either fluorescent or radioactive labeling

Since its invention in the early 1990s, differential display (DD) has become one of the most commonly used techniques for identifying differentially expressed genes at the mRNA level. Unlike other genomic approaches, such as DNA microarrays, DD systematically detects changes in mRNA profiles among multiple samples being compared without the need of any prior knowledge of genomic information of the living organism being studied. Here, we present an optimized DD protocol with a fluorescent digital readout as well as traditional radioactive labeling. The resulting streamlined fluorescent DD process offers an unprecedented accuracy, sensitivity and throughput in comprehensive and quantitative analysis of eukaryotic gene expression. Results usually can be obtained within days using a limited number of primer combinations, but a comprehensive DD screen may take weeks or months to accomplish, depending on gene coverage required and the number of differentially expressed genes present within a biological system being compared.     

Ecological footprint study on tourism itinerary products in Shangri-La, Yunnan Province, China

The eco-footprint analysis of tourism is one of the most up-to-date and effective methods used to analyze the environmental effects of tourism. This study constructs a model to calculate the ecological footprint (EF) of tourism itinerary products by using a component approach, rudimentarily exploring the calculation methods for EF which target necklace-like tourism itinerary products and base tourist trips. By applying the model to calculate and analyze an “8-day tour of Shangri-La”, a typical tourism itinerary product, results of this study suggest that: (a) Tourism is a kind of life style with tremendous ecological consumption, that is, per capita EF that tourists produce in the course of travel is more than the one that local people produce in their daily life in tourist source areas, and it also exceeds the per capita EF that local people produce in their daily life in tourist destination; (b) According to the component approach, EF of tourism itinerary products is broken down into 7 components, among which “Transport”, “Food”, “Waste” and “Accommodation” play important roles; (c) There exist significant differences in ecological efficiency between different departments of tourism; the travel and entertainment sectors maintain a relatively high ecological efficiency, while the food and lodging departments have relatively low ecological efficiency.     

Ecological application of wavelet analysis in the scaling of spatial distribution patterns of Ceratoides ewersmanniana

Ecological experiments are usually conducted on small scales, but the ecological and environmental issues are usually on large scales. Hence, there is a clear need of scaling. Namely, when we deal with patterns and processes on larger scales, a special connection needs to be established on the small scales that we are familiar with. Here we presented a wavelet analysis method that could build relationships between spatial distribution patterns on different scales. With this method, we also studied how spatial heterogeneity and distribution patterns changed with the scale. We investigated the distribution and the habitat of C. ewersmanniana in two plots (200 m × 200 m; the distance between these two plots is 15 km) at Mosuowan desert. The results demonstrated that spatial heterogeneity and distribution patterns were incorporated into larger scales when the wavelet scale varied from one (5 m) to four (20 m). However, if the wavelet scale was above five (25 m), the spatial distribution patterns varied placidly, the oscillation frequency of landforms stabilized at 110 m, and the dynamic quantity period of C. ewersmanniana stabilized at 115–125 m. We also identified signal mutation points with wavelet analysis and verified the heterogeneity degree of local space with position variance. We found that position variance decomposed the distribution patterns on large scales into small sampling plots, and the position with the largest variance also had the strongest heterogeneity. In a word, the wavelet analysis method could scale-up spatial distribution patterns and habitat heterogeneity. With this method and other methods derived from this one, such as wavelet scale, wavelet variance, position variance and extremely direct-viewing graphs, wavelet analysis could be widely applied in solving the scaling problem in ecological and environmental studies.     

Diversity of tropical forest landscape types in Hainan Island, China

A novel landscape classification system was proposed in this study based on landscape ecological theory and the differentiation in climate, topography, soil, vegetation and land use mode. Five basic units (zone, tract, province, region and type) and two assistant units (sub and group) were used in this system. The tropical forest landscape in Hainan Island was regarded as a landscape province, belonging to global tropical forest landscape zone, Asiatic (oriental) tropical forest landscape tract and Chinese tropical forest landscape subtract. Based on the grade system of region, sub-region, type-group and type, this landscape province (Hainan Island) is divided into 6 landscape regions (east moist forest landscape, west semi-arid forest landscape, central-south mountainous forest landscape, tropical evergreen needle-leaved forest landscape, tropical bamboo landscape and tropical plantation landscape), 11 tropical forest landscape sub-regions represented by tropical lowland valley rain forest landscape, 26 tropical forest landscape-type groups represented by tropical lowland valley Dipterocarpaceae forest landscape, and 54 forest landscape types (FLT) represented by lowland valley Vatica mangachapoi forest. Generally, this classification system represents the landscape diversity of the tropical forest in Hainan Island. Further studies are needed to better understand the landscape diversity of Hainan Island.     

A mutation in MRH2 kinesin enhances the root hair tip growth defect caused by constitutively activated ROP2 small GTPase in Arabidopsis

Root hair tip growth provides a unique model system for the study of plant cell polarity. Transgenic plants expressing constitutively active (CA) forms of ROP (Rho-of-plants) GTPases have been shown to cause the disruption of root hair polarity likely as a result of the alteration of actin filaments (AF) and microtubules (MT) organization. Towards understanding the mechanism by which ROP controls the cytoskeletal organization during root hair tip growth, we have screened for CA-rop2 suppressors or enhancers using CA1-1, a transgenic line that expresses CA-rop2 and shows only mild disruption of tip growth. Here, we report the characterization of a CA-rop2 enhancer (cae1-1 CA1-1) that exhibits bulbous root hairs. The cae1-1 mutation on its own caused a waving and branching root hair phenotype. CAE1 encodes the root hair growth-related, ARM domain-containing kinesin-like protein MRH2 (and thus cae1-1 was renamed to mrh2-3). Cortical MT displayed fragmentation and random orientation in mrh2 root hairs. Consistently, the MT-stabilizing drug taxol could partially rescue the wavy root hair phenotype of mrh2-3, and the MT-depolymerizing drug Oryzalin slightly enhanced the root hair tip growth defect in CA1-1. Interestingly, the addition of the actin-depolymerizing drug Latrunculin B further enhanced the Oryzalin effect. This indicates that the cross-talk of MT and AF organization is important for the mrh2-3 CA1-1 phenotype. Although we did not observe an apparent effect of the MRH2 mutation in AF organization, we found that mrh2-3 root hair growth was more sensitive to Latrunculin B. Moreover, an ARM domain-containing MRH2 fragment could bind to the polymerized actin in vitro. Therefore, our genetic analyses, together with cell biological and pharmacological evidence, suggest that the plant-specific kinesin-related protein MRH2 is an important component that controls MT organization and is likely involved in the ROP2 GTPase-controlled coordination of AF and MT during polarized growth of root hairs.