Influence of a short-term increase in intraluminal pressure, with and without recovery, on ACh-induced dilation in senescent skeletal muscle feed arteries.

Influence of a short-term increase in intraluminal pressure, with and without recovery, on ACh-induced dilation in senescent skeletal muscle feed arteries. John W. Seawright1 and Christopher R. Woodman1,2 1. Vascular Biology Laboratory, Dept. of Health and Kinesiology, Texas A&M University, College Station, TX 77843 2. Dept of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX 77843 We tested the hypothesis that a short-term increase in intraluminal pressure, to mimic a 1 h bout of exercise, would attenuate or reverse age-induced impairments in acetylcholine (ACh)-induced dilation in soleus muscle feed arteries (SFA). In addition, we hypothesized that improved endothelial function would persist following a 2 h recovery period at normal pressure. SFA were isolated from young (4 mo) and old (24 mo) Fischer 344 rats. SFA were cannulated and pressurized at 90 (p90) or 130 (p130) cm H2O for 1 h. At the end of the 1-h treatment period, p130 SFA were lowered to 90 cm H2O and ACh-induced vasodilation was assessed. In a separate group of SFA, pressure was raised to 130 cm H2O for 1 h and subsequently lowered to 90 cm H2O for a 2 h recovery period. ACh-induced vasodilator responses were significantly blunted in old p90 SFA relative to young p90 SFA. Pre-treatment with increased pressure (p130) for 1 h improved ACh-induced dilation in old (not young) SFA. The beneficial effect of pressure in old SFA was not apparent after a 2 h recovery period. To determine the importance of nitric oxide in endothelium-dependent dilation in pressure treated vessels, old SFA were pressurized at 130 cm H2O for 1 h, reset to p90, and ACh-induced vasodilator responses were assessed in the absence or presence of a NOS Inhibitor (L-NNA). Results indicate that the beneficial effect of pressure treatment on ACh-induced dilation was eliminated in the presence of L-NNA. Collectively, these results indicate that a short-term (1 h) increase in intraluminal pressure improves endothelium-dependent dilation in old SFA due to enhanced nitric oxide bioavailability. Contrary to our hypothesis, the beneficial effect of pressure did not persist following a 2 h recovery period. Research supported by AHA grant 0765043Y (CRW), AHA grant 4150031 (CRW), and a Sydney and J.L. Huffines Institute of Sports Medicine Graduate Student Research Grant (JWS). First author classification: Doctoral Studen

Radio-Adaptive Responses of Mouse Myocardiocytes

One of the most significant occupational hazards to an astronaut is the frequent exposure to radiation. Commonly associated with increased risk for cancer related morbidity and mortality, radiation is also known to increase the risk for cardiovascular related disorders including: pericarditis, hypertension, and heart failure. It is believed that these radiation-induced disorders are a result of abnormal tissue remodeling. It is unknown whether radiation exposure promotes remodeling through fibrotic changes alone or in combination with programmed cell death. Furthermore, it is not known whether it is possible to mitigate the hazardous effects of radiation exposure. As such, we assessed the expression and mechanisms of radiation-induced tissue remodeling and potential radio-adaptive responses of p53-mediated apoptosis and fibrosis pathways along with markers for oxidative stress and inflammation in mice myocardium. 7 week old, male, C57Bl/6 mice were exposed to 6Gy (H) or 5cGy followed 24hr later with 6Gy (LH) Cs-137 gamma radiation. Mice were sacrificed and their hearts extirpated 4, 24, or 72hr after final irradiation. Real Time - Polymerase Chain Reaction was used to evaluate target genes. Pro-apoptotic genes Bad and Bax, pro-cell survival genes Bcl2 and Bcl2l2, fibrosis gene Vegfa, and oxidative stress genes Sod2 and GPx4 showed a reduced fold regulation change (Bad,-6.18; Bax,-6.94; Bcl2,-5.09; Bcl2l2,-4.03; Vegfa, -11.84; Sod2,-5.97; GPx4*,-28.72; * = Bonferroni adjusted p-value . 0.003) 4hr after H, but not after 4hr LH when compared to control. Other p53-mediated apoptosis genes Casp3, Casp9, Trp53, and Myc exhibited down-regulation but did not achieve a notable level of significance 4hr after H. 24hr after H, genetic down-regulation was no longer present compared to 24hr control. These data suggest a general reduction in genetic expression 4hrs after a high dose of gamma radiation. However, pre-exposure to 5cGy gamma radiation appears to facilitate a radio-adaptive response that mitigates the reduction in genetic expression associated with single high-dose gamma radiation exposure. This radio-adaptive response may serve as a potential countermeasure to radiation-induced myocardial remodeling and preserve the cardiovascular health of astronauts; thus, reducing the risks of human space exploration

Mechanical Signals Promoting Healthy Vascular Endothelial and Smooth Muscle Cell Function in Aged Arteries

Peripheral arterial function is impaired with age, increasing the risk of cardiovascular disease. One therapeutic modality for improving arterial function in aged arteries is aerobic exercise; however, the signals associated with exercise that lead to improved arterial function are not fully understood. The aim of the research in this dissertation is to determine the importance of hemodynamic changes associated with aerobic exercise (increased intraluminal pressure and shear stress) which may serve as mechanical signals to promote healthy vascular endothelial and smooth muscle function in aged arteries. We hypothesized that a short-duration increase in intraluminal pressure would: 1) improve nitric oxide (NO)-mediated endothelium-dependent dilation in aged soleus muscle feed arteries (SFA); 2) improve vasoconstrictor responses in aged SFA via the Rho pathway; 3) improve vascular smooth muscle contractility in aged denuded (endothelium removed) SFA through the Rho pathway; and, 4) in combination with increased shear stress, induce greater improvements of endothelium-dependent dilation than a short-duration increase in pressure alone. SFA from young (4 mo) and old (24 mo) Fischer 344 rats were isolated and cannulated. Intact SFA or denuded (endothelium removed) SFA were exposed to increased intraluminal pressure for 1 h before assessment of vasodilator or vasoconstrictor function. The results from these experiments demonstrated that exposure to a short-duration increase in pressure, within a range believed to be present in SFA during exercise 1) improved NO-mediated, endothelium-dependent vasodilator responses in aged SFA; 2) improved vasoconstrictor responses in aged intact SFA via the Rho pathway; 3) impaired vasoconstrictor responses in aged denuded SFA; and, 4) did not interact with increased shear stress to produce greater improvements in endothelium-dependent dilation than pressure alone. Collectively, these data indicate that a short-duration increase in intraluminal pressure improves age-impaired arterial function and suggests that increased intraluminal pressure is one mechanical signal associated with aerobic exercise which promotes healthy endothelial and smooth muscle function in aged arteries

SNP-induced Dilation Following a Short-term Intraluminal Pressure Increase in Aged Skeletal Muscle Feed Arteries

A decline in peripheral vascular function occurs with age and is associated with an increased risk for cardiovascular disease. Moderate, aerobic exercise has been documented to improve or reverse the age-related decline in vascular function. The signal(s) associated with exercise responsible for the improvement in vascular function is not known; however, increased intraluminal pressure that occurs during a bout of exercise has been proposed to play an integral role. Previously, we reported that exposure to a short-term increase in intraluminal pressure improves endothelium-dependent vasodilator responses in aged arteries. The purpose of this study was to determine whether pressure treatment enhances endothelium-independent vasodilator responses. We hypothesized that a short-term (1 h) increase in intraluminal pressure, to mimic the intraluminal pressure believed to be present during a 1 h bout of exercise, would not improve endothelium-independent vasodilator responses. Soleus muscle feed arteries from young (4 month) and old (24 month) Fischer 344 rats were isolated, cannulated, and pressurized at 90 (P90) or 130 (P130) cmH20 for 1 h. Following the 1 h pressure treatment, intraluminal pressure was lowered to 90 cmH20 for all vessels and SNP-induced vasodilation was assessed immediately or following a 2 h recovery period. SNP-induced dilation was significantly blunted in old P90 SFA when compared to young P90 SFA. Pre-treatment with increased intraluminal pressure for 1 h improved SNP-induced vasodilator responses and abolished the age group difference between the old and young SFA. Maximal dilation to SNP was greater in the young P130 compared to the young P90 SFA. The effects of the 1 h increased intraluminal pressure treatment were no longer present following a 2 h recovery period. Collectively, these results indicate that treatment of aged arteries with a short-term increase in intraluminal pressure to mimic pressure during a bout of exercise improves endothelium-independent dilation in aged arteries. In addition, these results suggest that a short-term increase in intraluminal pressure, associated with a single bout of exercise, is one signal contributing to the beneficial effect of exercise on vasodilator responses in aged arteries

Induction of a Radio-Adaptive Response by Low-dose Gamma Irradiation in Mouse Cardiomyocytes

One of the most significant occupational hazards to an astronaut is the frequent exposure to radiation. Commonly associated with increased risk for cancer related morbidity and mortality, radiation is also known to increase the risk for cardiovascular related disorders including: pericarditis, hypertension, and heart failure. It is believed that these radiation-induced disorders are a result of abnormal tissue remodeling. It is unknown whether radiation exposure promotes remodeling through fibrotic changes alone or in combination with programmed cell death. Furthermore, it is not known whether it is possible to mitigate the hazardous effects of radiation exposure. As such, we assessed the expression and mechanisms of radiation-induced tissue remodeling and potential radio-adaptive responses of p53-mediated apoptosis and fibrosis pathways along with markers for oxidative stress and inflammation in mice myocardium. 7 week old, male, C57Bl/6 mice were exposed to 6Gy (H) or 5cGy followed 24hr later with 6Gy (LH) 137Cs gamma radiation. Mice were sacrificed and their hearts extirpated 4, 24, or 72hr after final irradiation. Real Time - Polymerase Chain Reaction was used to evaluate target genes. Apoptotic genes Bad and Bax, pro-cell survival genes Bcl2 and Bcl2l2, fibrosis gene Vegfa, and oxidative stress genes Sod2 and GPx4 showed a reduced fold regulation change (Bad,-6.18; Bax,-6.94; Bcl2,-5.09; Bcl2l2,-4.03; Vegfa, -11.84; Sod2,-5.97; GPx4*,-28.72; * = Bonferroni adjusted p-value < or = 0.003) 4hr after H, but not after 4hr LH compared to control. Other p53-mediated apoptosis genes Casp3, Casp9, Trp53, and Myc exhibited down-regulation but did not achieve a notable level of significance 4hr after H. 24hr after H, genetic down-regulation was no longer present compared to 24hr control. These data suggest a general reduction in genetic expression 4hrs after a high dose of gamma radiation. However, pre-exposure to 5cGy gamma radiation appears to facilitate a radio-adaptive response that mitigates the reduction in genetic expression associated with single high-dose gamma radiation exposure. This radio-adaptive response may serve as a potential countermeasure to radiation-induced myocardial remodeling and preserve the cardiovascular health of astronauts; thus, reducing the risks of human space exploration

Aging Impairs ACh-induced Dilation in Skeletal Muscle Feed Arteries: Role of Akt-dependent Phosphorylation of eNOS.

Aging impairs ACh-induced dilation in skeletal muscle feed arteries: role of Akt-dependent phosphorylation of eNOS. Meredith J. Luttrell1, John W. Seawright1, Daniel W. Trott1 and Christopher R. Woodman1,2 1. Dept. of Health and Kinesiology Texas A&M University, College Station, TX 77843 2. Dept of Veterinary Physiology and Pharmacology Texas A&M University, College Station, TX 77843 We tested the hypothesis that impaired nitric oxide (NO)-mediated, endothelium-dependent dilation in aged soleus muscle feed arteries (SFA) is due to an age-related decrement in PI3-kinase(PI3K)/protein kinase B (Akt)-dependent phosphorylation of endothelial NO synthase (eNOS) on serine residue 1177 (p-eNOSser1177). SFA from young (4 mo) and old (24 mo) Fischer 344 rats were cannulated for examination of endothelium-dependent vasodilator responses to acetylcholine (ACh). To determine the mechanism by which aging affected vasodilation to ACh, vasodilator responses were assessed in the absence and presence of Nw-nitro-L-arginine (L-NNA, to inhibit NOS), LY-294002 (to inhibit PI3K), or 1L6-hydroxymethyl-chiro-inositol-2-(R)-2-O-methyl-3-O-octadecyl-sn-glycerocarbonate (AktI, to inhibit Akt). Results indicate that ACh-induced vasodilation was significantly blunted in old SFA, whereas dilation to sodium nitroprusside (a NO donor) was not compromised. The age-group difference in ACh-induced dilation was abolished in the presence of L-NNA, LY-294002, or AktI. In a separate set of experiments, ACh-induced vasodilation was assessed in SFA from young and old rats. SFA were subsequently removed from the pipettes, snap frozen, and immunoblot analysis was used to assess p-AktSer473, p-eNOSser1177, total Akt and total eNOS protein content. ACh-induced vasodilation was blunted in old SFA; however, the p-AktSer473/Akt and p-eNOSser1177/eNOS ratios were similar in young and old SFA. Collectively, these results indicate that NO-mediated dilation is impaired in old SFA; however, the decrement in endothelial function is not due to reduced PI3K/Akt-dependent phosphorylation of eNOS on serine residue 1177

Biological Control of Giant Reed (Arundo donax): Economic Aspects

Arundo donax is a large, invasive weed consuming large quantities of water in the riparian area of the Texas Rio Grande Basin. With water availability a concern to the area, the USDA-ARS is investigating biological control agents to increase available water, creating a benefit to both the region’s economy and society in general.Arundo donax, Giant Reed, Water, Economics, Invasive, Environmental Economics and Policy, Resource /Energy Economics and Policy,

The Effect of Age and Exercise Training on Endothelial Function and Protein:Protein Interactions Among eNOS and Its Regulatory Proteins in Rat Aortas

Previous studies indicate that nitric oxide (NO)-mediated endothelium-dependent dilation declines with age. PURPOSE: The purpose of this study was to test the hypothesis that impaired NO-mediated, endothelium-dependent dilation in aged arteries is due to age-related alterations in protein:protein interactions among endothelial nitric oxide synthase (eNOS) and its regulatory proteins, resulting in reduced NO production in aged arteries. The regulatory proteins of interest in this study are Caveolin-1 (Cav1), an inhibitor of eNOS enzyme activity, and Heat-shock protein 90 (Hsp90), which enhances eNOS activity. METHODS: Young (2 mo) and Old (22 mo) male Fischer 344 rats were endurance exercise trained on a motorized treadmill or remained sedentary for 10wks yielding four groups of rats: 1)Young Sed (4 mo; n = 10), 2)Young Ex (4 mo; n = 10), 3) Old Sed (24 mo; n = 10), and 4) Old Ex (24 mo; n = 10). After the 10 week training period, rats were anesthetized; aortas were removed, cut into rings, and mounted on a wire myograph for assessment of endothelial function. Endothelium-dependent relaxation was assessed in aortic rings using acetylcholine (ACh). Endothelium-independent relaxation was assessed with sodium nitroprusside (SNP). Additional segments of aortas were isolated and snap frozen for use in co-immunoprecipitation experiments to assess Cav1:eNOS and Hsp90:eNOS protein:protein interactions. RESULTS: ACh-induced relaxation was impaired in aged aortic rings. Exercise training improved endothelium-dependent relaxation in old aortic rings such that ACh-induced relaxation in Old Ex aortas did not differ from Young Sed. Results from co-immunoprecipitation experiments revealed no significant age differences in the Cav1:eNOS interactions. Additionally, there was no significant training effect on Cav1:eNOS interaction in either young or old aortas. However, the Hsp90:eNOS interaction revealed a trend toward reduced interaction between the two proteins in the Old Sed group compared to the Young Sed group (p = 0.087). There was no significant training effect on Hsp90:eNOS interaction in young or old aortas. CONCLUSION: Ten weeks of exercise training improves endothelium-dependent relaxation in aortas from old rats. This improvement does not appear to be due to exercise-induced alterations in protein:protein interactions among eNOS and its regulatory proteins

Economic Implications of Biological Control of Arundo donax in the Texas Rio Grande Basin

ABSTRACT Arundo donax, or giant reed, is a large, bamboo-like plant that is native to Spain and has invaded several thousand acres of the Rio Grande riparian zone in Texas and Mexico. The plant grows to over 26 feet tall, and consumes large quantities of water, estimated as an amount equivalent to about 11% of irrigation water diverted by Valley irrigation districts (i.e., some estimates are more than 5.5 acre-feet per acre). With concern of increased water demands in the Texas Lower Rio Grande Valley region, the United States Department of Agriculture, Agricultural Research Service (USDA)ARS) is investigating four herbivorous insects as potential biological control agents for Arundo donax to facilitate increased water supply. This study examines selected economic implications for agricultural water users in the United States of applying these biological control agents along the Rio Grande. The research includes (a) estimating the value of the water saved due to the reduction of Arundo donax, (b) a benefit -cost analyses, (c) regional economic impact analyses, and (d) an estimate of the per-unit cost of water saved over a 50-year planning horizon (2009 through 2058). The model ArundoEcon© is used to perform a baseline deterministic analyses using low- and high-value irrigated composite acre values. That is, the saved water is initially valued based on being applied to agriculture as irrigation. Since the actual crop mix irrigated with the saved water is unknown, a range is provided by assuming all irrigated crops are “low-value,” and then again by including both “lowvalue” and “high-value” irrigated crops. Results of the water amount saved are 2/9 of the amount consumed, or approximately one acrefoot of water for each acre of Arundo. For each acre-foot of water saved, 1.85 dryland acres can be converted to low-value crop acres, and 0.71 can be converted to high-value crop acres. Regional economic results indicate a present value of farm-level benefits ranging from $98 to$160 million. Benefit-cost ratios are calculated with normalized prices and indicate a range from 4.38 to 8.81. Sensitivity analyses provide a robust set of results for Arundo agricultural water use, effectiveness of control agents, replacement species’ water use, Arundo expansion rate after control, value of water, and the cost of the program. The pre-production processes and farm-gate economic impact analyses are estimated using multipliers from the IMPLAN model. Regional results reveal a range of $9 to$18 million annually in economic output and 197 to 351 jobs associated with the increase in gross revenues due to the control of Arundo donax for the year 2025. Values for other select years are also provided. Further results suggest a life-cycle cost per acre-foot of water saved of $44. This amount is comparable to other projects designed to conserve water in the region. The USDA)ARS, Weslaco, Texas Arundo donax biological control project will realize positive results as indicated by the benefit-cost ratios, economic impact analyses, and competitive results for the per-unit cost of saving water. These results indicate this project will have positive economic implications for the U.S. and the Texas Lower Rio Grande Valley

Long-range downstream enhancers are essential for Pax6 expression

AbstractPax6 is a developmental control gene with an essential role in development of the eye, brain and pancreas. Pax6, as many other developmental regulators, depends on a substantial number of cis-regulatory elements in addition to its promoters for correct spatiotemporal and quantitative expression. Here we report on our analysis of a set of mice transgenic for a modified yeast artificial chromosome carrying the human PAX6 locus. In this 420 kb YAC a tauGFP-IRES-Neomycin reporter cassette has been inserted into the PAX6 translational start site in exon 4. The YAC has been further engineered to insert LoxP sites flanking a 35 kb long, distant downstream regulatory region (DRR) containing previously described DNaseI hypersensitive sites, to allow direct comparison between the presence or absence of this region in the same genomic context. Five independent transgenic lines were obtained that vary in the extent of downstream PAX6 locus that has integrated. Analysis of transgenic embryos carrying full-length and truncated versions of the YAC indicates the location and putative function of several novel tissue-specific enhancers. Absence of these distal regulatory elements abolishes expression in specific tissues despite the presence of more proximal enhancers with overlapping specificity, strongly suggesting interaction between these control elements. Using plasmid-based reporter transgenic analysis we provide detailed characterization of one of these enhancers in isolation. Furthermore, we show that overexpression of a short PAX6 isoform derived from an internal promoter in a multicopy YAC transgenic line results in a microphthalmia phenotype. Finally, direct comparison of a single-copy line with the floxed DRR before and after Cre-mediated deletion demonstrates unequivocally the essential role of these long-range control elements for PAX6 expression