Guanylate cyclase C limits systemic dissemination of a murine enteric pathogen

BACKGROUND: Guanylate Cyclase C (GC-C) is an apically-oriented transmembrane receptor that is expressed on epithelial cells of the intestine. Activation of GC-C by the endogenous ligands guanylin or uroguanylin elevates intracellular cGMP and is implicated in intestinal ion secretion, cell proliferation, apoptosis, intestinal barrier function, as well as the susceptibility of the intestine to inflammation. Our aim was to determine if GC-C is required for host defense during infection by the murine enteric pathogen Citrobacter rodentium of the family Enterobacteriacea. METHODS: GC-C(+/+) control mice or those having GC-C genetically ablated (GC-C(−/−)) were administered C. rodentium by orogastric gavage and analyzed at multiple time points up to post-infection day 20. Commensal bacteria were characterized in uninfected GC-C(+/+) and GC-C(−/−) mice using 16S rRNA PCR analysis. RESULTS: GC-C(−/−) mice had an increase in C. rodentium bacterial load in stool relative to GC-C(+/+). C. rodentium infection strongly decreased guanylin expression in GC-C(+/+) mice and, to an even greater degree, in GC-C(−/−) animals. Fluorescent tracer studies indicated that mice lacking GC-C, unlike GC-C(+/+) animals, had a substantial loss of intestinal barrier function early in the course of infection. Epithelial cell apoptosis was significantly increased in GC-C(−/−) mice following 10 days of infection and this was associated with increased frequency and numbers of C. rodentium translocation out of the intestine. Infection led to significant liver histopathology in GC-C(−/−) mice as well as lymphocyte infiltration and elevated cytokine and chemokine expression. Relative to naïve GC-C(+/+) mice, the commensal microflora load in uninfected GC-C(−/−) mice was decreased and bacterial composition was imbalanced and included outgrowth of the Enterobacteriacea family. CONCLUSIONS: This work demonstrates the novel finding that GC-C signaling is an essential component of host defense during murine enteric infection by reducing bacterial load and preventing systemic dissemination of attaching/effacing-lesion forming bacterial pathogens such as C. rodentium

The social networks of manureshed management

Manureshed management—the strategic use of manure nutrients that prioritizes recycling between livestock systems and cropping systems—provides a comprehensive framework for sustainable nutrient management that necessitates the collaboration of many actors. Understanding the social dimensions of collaboration is critical to implement the strategic and technological requirements of functional manuresheds. To improve this understanding, we identified aspirational networks of actors involved in manureshed management across local, regional, and national scales, principally in the United States, elucidating key relationships and highlighting the breadth of interactions essential to successful manureshed management. We concluded that, although the social networks vary with scale, the involvement of a common core set of actors and relationships appears to be universal to the successful integration of modern livestock and crop production systems necessary for functional manuresheds. Our analysis also reveals that, in addition to agricultural producers, local actors in extension and advisory services and private and public sectors ensure optimal outcomes at all scales. For manureshed management to successfully integrate crop and livestock production and sustainably manage manure nutrient resources at each scale, the full complement of actors identified in these social networks is critical to generate innovation and ensure collaboration continuity

Prognostic value of stress hyperglycemia ratio on short- and long-term mortality after acute myocardial infarction

Aims: Prior studies demonstrated an association between hospital admission blood glucose and mortality in acute myocardial infarction (AMI). Because stress hyperglycemia ratio (SHR) has been suggested as a more reliable marker of stress hyperglycemia this study investigated to what extent SHR in comparison with admission blood glucose is associated with short- and long-term mortality in diabetic and non-diabetic AMI patients. Methods: The analysis was based on 2,311 AMI patients aged 25–84 years from the population-based Myocardial Infarction Registry Augsburg (median follow-up time 6.5 years [IQR: 4.9–8.1]). The SHR was calculated as admission glucose (mg/dl)/(28.7 × HbA1c (%)—46.7). Using logistic and COX regression analyses the associations between SHR and admission glucose and mortality were investigated. Result: Higher admission glucose and higher SHR were significantly and nonlinearly associated with higher 28-day mortality in AMI patients with and without diabetes. In patients without diabetes, the AUC for SHR was significantly lower than for admission glucose (SHR: 0.6912 [95%CI 0.6317–0.7496], admission glucose: 0.716 [95%CI 0.6572–0.7736], p-value: 0.0351). In patients with diabetes the AUCs were similar for SHR and admission glucose. Increasing admission glucose and SHR were significantly nonlinearly associated with higher 5-year all-cause mortality in AMI patients with diabetes but not in non-diabetic patients. AUC values indicated a comparable prediction of 5-year mortality for both measures in diabetic and non-diabetic patients. Conclusions: Stress hyperglycemia in AMI patients plays a significant role mainly with regard to short-term prognosis, but barely so for long-term prognosis, underlining the assumption that it is a transient dynamic disorder that occurs to varying degrees during the acute event, thereby affecting prognosis

The inositol pyrophosphate metabolism of Dictyostelium discoideum does not regulate inorganic polyphosphate (polyP) synthesis

Initial studies on the inositol phosphates metabolism were enabled by the social amoeba Dictyostelium discoideum. The abundant amount of inositol hexakisphosphate (IP6 also known as Phytic acid) present in the amoeba allowed the discovery of the more polar inositol pyrophosphates, IP7 and IP8, possessing one or two high energy phosphoanhydride bonds, respectively. Considering the contemporary growing interest in inositol pyrophosphates, it is surprising that in recent years D. discoideum, has contributed little to our understanding of their metabolism and function. This work fulfils this lacuna, by analysing the ip6k, ppip5k and ip6k-ppip5K amoeba null strains using PAGE, 13C-NMR and CE-MS analysis. Our study reveals an inositol pyrophosphate metabolism more complex than previously thought. The amoeba Ip6k synthesizes the 4/6-IP7 in contrast to the 5-IP7 isomer synthesized by the mammalian homologue. The amoeba Ppip5k synthesizes the same 1/3-IP7 as the mammalian enzyme. In D. discoideum, the ip6k strain possesses residual amounts of IP7. The residual IP7 is also present in the ip6k-ppip5K strain, while the ppip5k single mutant shows a decrease in both IP7 and IP8 levels. This phenotype is in contrast to the increase in IP7 observable in the yeast vip1Δ strain. The presence of IP8 in ppip5k and the presence of IP7 in ip6k-ppip5K indicate the existence of an additional inositol pyrophosphate synthesizing enzyme. Additionally, we investigated the existence of a metabolic relationship between inositol pyrophosphate synthesis and inorganic polyphosphate (polyP) metabolism as observed in yeast. These studies reveal that contrary to the yeast, Ip6k and Ppip5k do not control polyP cellular level in amoeba

Environmental Management of Grazing Lands

Bacteria levels are the number one cause of water quality impairment in Texas. Several recent Total Maximum Daily Loads (TMDLs) in Texas, such as those implemented in the Peach Creek and Leon River watersheds, have identified grazing cattle as a contributor to bacterial water quality impairments in those watersheds through both direct deposition and runoff of their fecal matter to streams. To address this issue, the Texas State Soil and Water Conservation Board (TSSWCB) and the Natural Resources Conservation Service (NRCS) funded this project to assist with development and delivery of technical information and support to ranchers on protection and enhancement of the functions and values of grasslands. A number of best management practices (BMPs) have been identified to reduce bacteria runoff from grazing lands and direct deposition into streams. The primary focus of these BMPs is to maintain adequate ground cover and minimize concentrated livestock areas, especially on sensitive areas such as riparian areas. Maintaining adequate ground cover and plant density improves the filtering capacity of the vegetation and enhances water infiltration into the soil. Minimizing concentrated livestock areas, trailing, and trampling reduces soil compaction, reduces excess runoff and subsequent soil erosion, and enhances fecal matter distribution and ground cover. Specific BMPs identified include grazing management, fencing, alternate water sources, hardened watering points, controlled access, supplemental feed placement, and shade or cover manipulation (NRCS 2007). This project accomplished several objectives, including: 1) compiling existing information on environmental management of grazing lands, 2) evaluating and demonstrating the effectiveness of proper grazing management in reducing bacterial runoff from grazing lands, 3) initiating evaluation of the effect of complementary practices (i.e. alternative water supplies and shade) on cattle behavior and stream water quality, and 4) promoting adoption of appropriate grazing land management practices. Evaluation and demonstration of the effect of grazing management on bacteria runoff at the USDA-ARS Riesel Watersheds has produced some interesting results. The site mean concentration of E. coli (i.e. flow weighted concentration) at the ungrazed native prairie site was surprisingly high (1.0E+04 cfu/100 ml), greatly exceeding the Texas Surface Water Quality Standards single sample standard for E. coli (394 cfu/100 ml) as well as the geometric mean (126 cfu/100 ml). It is important to note that these standards apply to waterbodies, such as streams and reservoirs, but not to edge-of-field runoff as described here. Also, the E. coli concentration seen in the runoff from the moderately grazed bermudagrass site (2.3E+04 cfu/100 ml) was significantly higher (more than double) than that observed at the native prairie site. These levels, however, are consistent with the findings of other researchers. The pre-BMP implementation evaluation of the effectiveness of alternative water supplies and shade on cattle behavior and stream water quality (E. coli) has been completed at the 2S Ranch, near Lockhart. This evaluation showed that when alternative water was not available, E. coli levels increased as the stream flowed through the ranch. Quarterly evaluation of cattle behavior using GPS collars indicated cattle spent only 4.5% of the time within 35 feet of the stream when alternative water was not available. When alternative water was provided, however, this percent time that cattle spent within 35 feet of the stream was reduced to 1.1%, a 75% reduction. This reduction is consistent with the findings of other researchers. Post-BMP evaluation has been initiated and will continue for another year now that alternative water and shade has been provided. Much was done through this project to increase awareness of the bacteria issue and BMPs to address them. AgriLife Extension and TWRI developed fact sheets, provided posters and presentations, conducted site tours, and developed a Web site to help disseminate information. These outreach activities reached local, state, and national audiences. The Web site alone has reached 539 unique visitors during the project. Much is left to do, however. Evaluation of grazing management, alternative water supplies, and shade will continue. Data on other practices (i.e. using rip-rap to reduce access to riparian areas, providing controlled access points, etc.) and groups of practices is still needed to provide cattlemen with a “toolbox” for addressing the bacteria issue. Modification of water quality standards may also be appropriate to address the high levels of E. coli found in runoff from ungrazed sites. Education programs need to be conducted state- and nation-wide to assist cattlemen in addressing bacteria issues

Inositol pyrophosphates promote the interaction of SPX domains with the coiled-coil motif of PHR transcription factors to regulate plant phosphate homeostasis

Phosphorus is an essential nutrient taken up by organisms in the form of inorganic phosphate (Pi). Eukaryotes have evolved sophisticated Pi sensing and signaling cascades, enabling them to stably maintain cellular Pi concentrations. Pi homeostasis is regulated by inositol pyrophosphate signaling molecules (PP-InsPs), which are sensed by SPX domain-containing proteins. In plants, PP-InsP-bound SPX receptors inactivate Myb coiled-coil (MYB-CC) Pi starvation response transcription factors (PHRs) by an unknown mechanism. Here we report that a InsP8–SPX complex targets the plant-unique CC domain of PHRs. Crystal structures of the CC domain reveal an unusual four-stranded anti-parallel arrangement. Interface mutations in the CC domain yield monomeric PHR1, which is no longer able to bind DNA with high affinity. Mutation of conserved basic residues located at the surface of the CC domain disrupt interaction with the SPX receptor in vitro and in planta, resulting in constitutive Pi starvation responses. Together, our findings suggest that InsP8 regulates plant Pi homeostasis by controlling the oligomeric state and hence the promoter binding capability of PHRs via their SPX receptors

Harnessing {13}^C-labeled myo-inositol to interrogate inositol phosphate messengers by NMR

Inositol poly- and pyrophosphates (InsPs and PP-InsPs) are an important group of metabolites and mediate a wide range of processes in eukaryotic cells. To elucidate the functions of these molecules, robust techniques for the characterization of inositol phosphate metabolism are required, both at the biochemical and the cellular level. Here, a new tool-set is reported, which employs uniformly {13}^C-labeled compounds ([{13}^C_{6}] myo-inositol, ([{13}^C_{6}]InsP_{5}, ([{13}^C_{6}]InsP_{6}, and ([{13}^C_{6}]5PP-InsP_{5}), in combination with commonly accessible NMR technology. This approach permitted the detection and quantification of InsPs and PP-InsPs within complex mixtures and at physiological concentrations. Specifically, the enzymatic activity of IP6K1 could be monitored in vitro in real time. Metabolic labeling of mammalian cells with [{13}^C_{6}]myo-inositol enabled the analysis of cellular pools of InsPs and PP-InsPs, and uncovered high concentrations of 5PP-InsP_{5} in HCT116 cells, especially in response to genetic and pharmacological perturbation. The reported method greatly facilitates the analysis of this otherwise spectroscopically silent group of molecules, and holds great promise to comprehensively analyze inositol-based signaling molecules under normal and pathological conditions