RESUMEN

The probiotic yeast Saccharomyces boulardii has great potential for use as a chassis for microbiome engineering because of its high resistance to environmental stress, well-developed genetic tools, and the ability to secrete recombinant proteins in the intestine. As oral feeding of lysozyme has been reported to change the gut microbiome and fecal metabolites, we engineered S. boulardii to secrete human lysozyme, and investigated the changes in the microbiome and fecal metabolites in response to the administration of the engineered probiotic yeast into mice. Administration of S. boulardii changed the structure of the gut microbiome by promoting the growth of clostridia and increasing the diversity of strains. The human lysozyme secreted by S. boulardii in the intestine resulted in a unique gut microbiome structure through selective growth. In addition, the administration of probiotic yeast S. boulardii affected host energy metabolism and decreased blood urea and fructose levels, suggesting a mechanism of health benefits in mice. IMPORTANCE Our study identified changes in the microbiome by administering wild-type S. boulardii in mice to healthy mice based on long-read sequencing and demonstrated that a recombinant protein secreted by engineered S. boulardii in the intestine could change the microbiome. Our results provide valuable information for the development of therapeutics using engineered S. boulardii that changes the gut microbiome and host physiology.

Asunto(s)

Microbioma Gastrointestinal , Microbiota , Probióticos , Saccharomyces boulardii , Humanos , Animales , Ratones , Saccharomyces boulardii/genética , Saccharomyces boulardii/metabolismo , Muramidasa/genética , Saccharomyces cerevisiae/metabolismo , Metaboloma

RESUMEN

Microalgae are promising sustainable resources because of their ability to convert CO2 into biofuels and chemicals directly. However, the industrial production and economic feasibility of microalgal bioproducts are still limited. As such, metabolic engineering approaches have been undertaken to enhance the productivities of microalgal bioproducts. In the last decade, impressive advances in microalgae metabolic engineering have been made by developing genetic engineering tools and multi-omics analysis. This review presents comprehensive microalgal metabolic pathways and metabolic engineering strategies for producing lipids, long chain-polyunsaturated fatty acids, terpenoids, and carotenoids. Additionally, promising metabolic engineering approaches specific to target products are summarized. Finally, this review discusses current challenges and provides future perspectives for the effective production of chemicals and fuels via microalgal metabolic engineering.

Asunto(s)

Microalgas , Biocombustibles , Lípidos , Ingeniería Metabólica , Redes y Vías Metabólicas

RESUMEN

The green alga Chlamydomonas reinhardtii serves as a model organism for plant and photosynthesis research due to many commonalities in metabolism and to the fast growth rate of C. reinhardtii which accelerates experimental turnaround time. In addition, C. reinhardtii is a focus of research efforts in metabolic engineering and synthetic biology for the potential production of biofuels and value-added chemicals. Here, we report that the C. reinhardtii cia5 mutant, which lacks a functional carbon-concentrating mechanism (CCM), can produce substantial amounts of glycolate, a high-value cosmetic ingredient, when the mutant is cultured under ambient air conditions. In order to reveal the metabolic basis of glycolate accumulation by the cia5 mutant, we investigated the metabolomes of the cia5 mutant and a wild type strain CC-125 (WT) through the global metabolic profiling of intracellular and extracellular fractions using gas chromatography and mass spectrometry. We observed the intracellular and extracellular metabolic profiles of the WT and the cia5 mutant were similar during the mixotrophic phase at 30 h. However, when the cells entered the photoautotrophic phase (i.e., 96 h and 120 h), both the intracellular and extracellular metabolic profiles of cia5 mutant differed significantly when compared to WT. In the cia5 mutant strain, a group of photorespiration pathway intermediates including glycolate, glyoxylate, glycine, and serine accumulated to significantly higher levels compared to WT. In the photorespiration pathway, glycolate is metabolized to glyoxylate and glycine leading to NH3 and CO2 generation during the mitochondrial conversion of glycine to serine. This result provides further evidence that the CIA5 mutation increased the photorespiration rate. Because the cia5 mutant lacks a CCM, and C. reinhardtii might harbor an inefficient or incomplete photorespiration pathway, glycolate may accumulate when the CCM is not functional. We envision that investigating photorespiration controls in C. reinhardtii provides tools for producers to use the cia5 mutant to produce glycolate as well as platform to engineer alternative pathways for glycolate metabolism.

Asunto(s)

Chlamydomonas reinhardtii , Carbono , Dióxido de Carbono , Chlamydomonas reinhardtii/genética , Cromatografía de Gases y Espectrometría de Masas , Glicolatos , Fotosíntesis/genética

RESUMEN

INTRODUCTION: Disturbances in the visual, vestibular, and oculomotor systems have been identified in Parkinson's disease (PD). Patients' perspectives regarding these symptoms remain unexplored and may provide insights on functional implications of these symptoms and guide future interventions. The goal of this study is to elicit perceptions of individuals with PD with respect to visual, vestibular, and oculomotor deficits. Methods: Twenty-nine individuals with PD participated in focus group discussions. Participants discussed visual, vestibular, and oculomotor deficits they experience and how these deficits affect function. Discussions were recorded, transcribed, and coded. Inductive qualitative data analysis techniques were used to interpret responses. Results: Four themes emerged: 1) participants perceived visual, vestibular, and oculomotor deficits and related these deficits to their PD diagnosis; 2) participants perceive that these deficits affect function; 3) participants suggested these deficits are not recognized by healthcare providers; and 4) participants indicated they receive limited treatment for these deficits. Conclusions: Visual, vestibular, and oculomotor deficits are under-reported and under-assessed symptoms, which have a significant impact on the lives of people with PD. Healthcare providers should be aware of such deficits. The findings suggest that the healthcare team can better identify these deficits and identify important future areas of research.

Asunto(s)

Trastornos de la Motilidad Ocular/fisiopatología , Enfermedad de Parkinson/fisiopatología , Vestíbulo del Laberinto/fisiopatología , Trastornos de la Visión/fisiopatología , Anciano , Femenino , Grupos Focales , Humanos , Masculino , Persona de Mediana Edad , Participación del Paciente , Investigación Cualitativa

RESUMEN

ß-Carotene is a natural pigment and health-promoting metabolite, and has been widely used in the nutraceutical, feed, and cosmetic industries. Here, we engineered a GRAS yeast Saccharomyces cerevisiae to produce ß-carotene from xylose, the second most abundant and inedible sugar component of lignocellulose biomass. Specifically, a ß-carotene biosynthetic pathway containing crtYB, crtI, and crtE from Xanthophyllomyces dendrorhous was introduced into a xylose-fermenting S. cerevisiae. The resulting strain produced ß-carotene from xylose at a titer threefold higher than from glucose. Interestingly, overexpression of tHMG1, which has been reported as a critical genetic perturbation to enhance metabolic fluxes in the mevalonate pathway and ß-carotene production in yeast when glucose is used, did not further improve the production of ß-carotene from xylose. Through fermentation profiling, metabolites analysis, and transcriptional studies, we found the advantages of using xylose as a carbon source, instead of glucose, for ß-carotene production to be a more respiratory feature of xylose consumption, a larger cytosolic acetyl-CoA pool, and an upregulated expression level of rate-limiting genes in the ß-carotene-producing pathway, including ACS1 and HMG1. As a result, 772.8 mg/L of ß-carotene was obtained in a fed-batch bioreactor culture with xylose feeding. Considering the inevitable large scale production of xylose when cellulosic biomass-based bioeconomy is implemented, our results suggest xylose utilization is a promising strategy for overproduction of carotenoids and other isoprenoids in engineered S. cerevisiae.

Asunto(s)

Ingeniería Metabólica/métodos , Saccharomyces cerevisiae , Xilosa/metabolismo , beta Caroteno/metabolismo , Basidiomycota/enzimología , Basidiomycota/genética , Reactores Biológicos/microbiología , Glucosa/metabolismo , Proteína HMGB1/genética , Proteína HMGB1/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo

RESUMEN

Chronic treatment with angiotensin receptor blockers is largely accepted for protecting cerebral circulation during hypertension, but beneficial effects of short-term treatments are questionable, as highlighted by the recent SCAST trial. We compared the impact of 10 days treatment with candesartan (as SCAST) versus telmisartan (previously described to reverse arteriolar remodeling, chronic treatment) on pial arterioles of spontaneously hypertensive rats (SHR). We explored whether PPAR-gamma agonist activity or AT(1) receptor blockade are involved in their differential effects. In the first study, 4-month-old male SHR were treated with telmisartan (TELMI, 2 mg/kg per day) or candesartan cilexetil (CANDE, 10 mg/kg per day) and compared to vehicle treated SHR and normotensive WKY. In a second study, SHR were treated with CANDE, pioglitazone (a PPAR-gamma agonist, PIO 2.5 mg/kg per day) or CANDE+PIO, compared to TELMI. Internal diameter of pial arterioles (ID, cranial window) was measured at baseline, during hemorrhage-induced hypotension, or following suffusion of Ang II (10(-6) mol/L) or EDTA inactivation of smooth muscle cells (passive ID). PPAR-gamma and eNOS (target gene of PPAR-gamma) mRNA were evaluated in brain microvessels. For similar antihypertensive effects, TELMI (+44% versus SHR), but not CANDE, increased baseline ID. During hemorrhage, ID in TELMI group was similar to WKY, while ID in SHR and CANDE remained lower. In the second study, TELMI (+36%, versus SHR) and CANDE+PIO (+43%) increased baseline ID, but not CANDE or PIO alone. TELMI (-66%) and CANDE+PIO (-69%), but neither CANDE nor PIO alone, decreased Ang II-induced vasoconstriction. CANDE+PIO, but not CANDE, increased passive ID. In both studies, PPAR-gamma and eNOS expressions were higher in TELMI than CANDE. Short-term treatment with TELMI, but not with CANDE, reverses narrowing of pial arteriolar ID in SHR. This may involve PPAR-gamma related mechanisms, since CANDE+PIO treatment induced similar effects, and a better blockade of AT(1) receptors.

Asunto(s)

Antagonistas de Receptores de Angiotensina/farmacología , Arteriolas/efectos de los fármacos , Circulación Cerebrovascular/efectos de los fármacos , Hipertensión/tratamiento farmacológico , Piamadre/irrigación sanguínea , Receptor de Angiotensina Tipo 1/química , Animales , Bencimidazoles/uso terapéutico , Benzoatos/uso terapéutico , Compuestos de Bifenilo , Encéfalo/metabolismo , Masculino , Microcirculación , Músculo Liso/efectos de los fármacos , Óxido Nítrico Sintasa de Tipo III/metabolismo , PPAR gamma/metabolismo , Pioglitazona , ARN Mensajero/metabolismo , Ratas , Ratas Endogámicas SHR , Telmisartán , Tetrazoles/uso terapéutico , Tiazolidinedionas/uso terapéutico

RESUMEN

BACKGROUND: Angiotensin II (Ang II) induces constriction (AT(1)) and dilation (AT(2) receptors) of cerebral arterioles. High sodium intake induces changes in receptors expression and loss of AT(2)-mediated vasodilation in extracerebral vessels. We investigated whether high salt modifies the AT(2)-mediated response of cerebral arterioles. METHODS: Three-month-old male Wistar rats received drinking water supplemented or not with 1% NaCl. We measured at day 4 or 30 plasma aldosterone concentration, AT receptors expression (brain microvessels, western blot, RT-qPCR), internal diameter of pial arterioles (cranial window) following suffusion with Ang II (10(-6) mol/l, or 10(-8) mol/l + losartan 10(-5) mol/l), serotonin (5-HT, 10(-6) mol/l), sodium nitroprusside (10(-5) mol/l) and adenosine diphosphate (ADP, 10(-4) mol/l). RESULTS: High salt did not modify arterial pressure, baseline arteriolar diameter, vasoconstriction to Ang II or 5-HT, nor vasodilation to SNP. High salt lowered plasma aldosterone concentration (d4 138 ± 71 not significant vs. control 338 ± 73; d30 150 ± 21 P < 0.05 vs. control 517 ± 79 µmol/l). AT receptors mRNA did not change while protein level of AT(2) receptors decreased at d4 (64 ± 9% of control, P < 0.05). AT(2)-mediated vasodilation (control d4; d30 8 ± 2; 5 ± 2%) was abolished at d4 (-2 ± 2%, P < 0.05) and reversed to vasoconstriction at d30 (-7 ± 2%, P < 0.05). ADP-induced vasodilation is abolished at d30 (2 ± 2, P < 0.05 vs. control 19 ± 4%). CONCLUSION: High salt specifically abolishes AT(2)-mediated vasodilation, immediately, via decreased level of AT(2) receptor protein, and after 30 days, in association with abolition of endothelial vasodilation. Such loss of AT(2)-mediated vasodilation may be deleterious in case of stroke.

Asunto(s)

Angiotensina II/fisiología , Arteriolas/fisiología , Cloruro de Sodio Dietético/administración & dosificación , Vasodilatación/fisiología , Animales , Secuencia de Bases , Cartilla de ADN , Masculino , Reacción en Cadena de la Polimerasa , Ratas , Ratas Wistar

RESUMEN

OBJECTIVE: Antihypertensive treatment with standard clinical doses of angiotensin-converting enzyme inhibitors (ACEIs) or angiotensin II receptor blockers (ARBs) reverses cerebral arteriolar remodeling, thus restoring dilatation and the lower limit of cerebral blood flow (CBF) autoregulation (LL CBF AR). In humans, a combination of standard clinical doses of the two drugs does not produce greater protection against stroke than that obtained with single-drug treatments and increases the risk of side-effects. We hypothesized that a combination of suboptimal doses of the ARB, telmisartan (TEL) and of the ACEI, ramipril (RAM), could be a well tolerated and effective treatment of hypertension-induced remodeling of cerebral arterioles. DESIGN: We studied the impact of 3-month oral treatment with TEL (0.5 or 0.8 mg/kg per day) or RAM (0.1 or 0.25mg/ kg per day) alone or in combination (TEL0.8 + RAM0.1 or TEL0.5 + RAM0.25) on the cerebral circulation of the spontaneously hypertensive rats (SHRs). Normotensive Wistar-Kyoto rats (WKYs) were taken as controls. METHODS: Cerebral arteriolar pressure, CBF and internal diameter were measured via an open-skull preparation at baseline and during hypotension before and after deactivation (EDTA). RESULTS: Combinations normalized cerebral arteriolar pressure, whereas drugs alone had no significant impact. TEL0.8 + RAM0.1 showed the greatest effect on arteriolar internal diameter (SHRs 42+/-16, WKYs 59+/-16microm, TEL0.5 + RAM0.25 50+/-6, TEL0.8 + RAM0.1 62+/-18, P<0.05) and normalized LL CBF AR (SHRs 77+/-28, WKYs 53+/-17 mmHg, TEL0.8 + RAM0.1 50+/-10, P<0.05). CONCLUSION: The combination of suboptimal doses of TEL and RAM with an 8 : 1 ratio has the greatest effect on cerebral circulation and could represent well tolerated and efficient treatment of cerebral ischemia and stroke.

Asunto(s)

Bloqueadores del Receptor Tipo 1 de Angiotensina II/administración & dosificación , Inhibidores de la Enzima Convertidora de Angiotensina/administración & dosificación , Inhibidores de la Enzima Convertidora de Angiotensina/farmacología , Arteriolas/efectos de los fármacos , Bencimidazoles/administración & dosificación , Bencimidazoles/farmacología , Benzoatos/administración & dosificación , Benzoatos/farmacología , Circulación Cerebrovascular/efectos de los fármacos , Hipertensión/tratamiento farmacológico , Ramipril/administración & dosificación , Ramipril/farmacología , Animales , Antihipertensivos/farmacología , Antihipertensivos/uso terapéutico , Arteriolas/fisiopatología , Presión Sanguínea/efectos de los fármacos , Quimioterapia Combinada , Hipertensión/fisiopatología , Masculino , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Telmisartán

RESUMEN

Moderate consumption of red wine is associated with a lower incidence of cardiovascular disease. Red wine polyphenols (RWP) have been proposed to be beneficial, but there is lack of evidence concerning the cerebrovascular effects of RWP. We studied the effect of local administration of a RWP extract (10(-2) mg/mL) on the diameter of rat cerebral arterioles using an open cranial window technique in vivo. We measured cerebral arteriolar diameter and systemic blood pressure. Cerebral arterioles reacted concentration-dependently to adenosine diphosphate [ADP, dilatation EC50 5.3 x 10(-5) M (95% confidence interval: 3.1 to 9.0 x 10 M(-5))], NG-nitro-L-arginine methyl ester [L-NAME, constriction EC50 5.8 x 10(-9) M (95% CI: 2.5 x 10(-9) to 1.4 x 10(-8) M)]. and sodium nitroprusside [SNP, dilatation EC50 1.0 x 10(-6) M (95% CI: 9.2 x 10(-7) to 1.1 x 10(-6) M)]. RWP enhanced vasodilation induced by ADP (10(-4) M) from 17 +/- 2% to 29 +/- 4% and reversed L-NAME-induced vasoconstriction but did not affect SNP-induced vasodilation. Systemic hypotension induced by hemorrhage caused myogenic arteriolar dilation. RWP further dilated cerebral arterioles (from -1 +/- 2% to 8 +/- 3%) with 1 mL of blood withdrawn. In summary, RWP improved endothelium-dependent and pressure-induced vasodilation in rat cerebral arterioles. This could be beneficial in improving cerebral blood flow under ischemic condition.

Asunto(s)

Cerebro/irrigación sanguínea , Endotelio Vascular/fisiología , Flavonoides/farmacología , Fenoles/farmacología , Vasodilatación/efectos de los fármacos , Vino , Adenosina Difosfato/farmacología , Animales , Arteriolas/fisiología , Relación Dosis-Respuesta a Droga , Masculino , NG-Nitroarginina Metil Éster/farmacología , Nitroprusiato/farmacología , Polifenoles , Ratas , Ratas Wistar

RESUMEN

Several reports suggest that consumption of red wine is associated with a lower risk of stroke. We investigated the chronic effect of red wine polyphenols (RWP) on the functional and structural characteristics of cerebral arterioles in chronic hypertension, which is an important risk factor of stroke. Spontaneous hypertensive rats (SHR) were treated with RWP extract (100 mg/kg/day in drinking water) for 10 weeks. We measured the effect of agonist- and hypotension-induced changes in internal diameter of cerebral arterioles using an open cranial window technique. Wall mechanical parameters were determined in deactivated cerebral arterioles. The activity of antioxidant enzymes in plasma was determined. Adenosine diphosphate-induced vasodilatation was decreased by 48% in SHR and normalized in SHR treated with RWP. RWP had no effect on hypotension-induced dilatation. RWP decreased the wall thickness/external diameter ratio by 13% and significantly shifted the stress-strain relationship of the arteriole wall to the left. There was a decrease in glutathione-S-transferase and glutathione peroxidase after treatment of RWP in SHR. In summary, chronic oral administration of RWP to SHR improved endothelium-dependent dilatation, normalized wall stress and diameter, and altered the systemic antioxidant state. These effects of RWP could be useful in the prevention of stroke in hypertensive patients.

Asunto(s)

Circulación Cerebrovascular/efectos de los fármacos , Hipertensión/tratamiento farmacológico , Fenoles/farmacología , Vino , Animales , Arteriolas/efectos de los fármacos , Arteriolas/fisiopatología , Modelos Animales de Enfermedad , Esquema de Medicación , Glutatión Peroxidasa/efectos de los fármacos , Glutatión Peroxidasa/metabolismo , Glutatión Transferasa/efectos de los fármacos , Glutatión Transferasa/metabolismo , Hipertensión/fisiopatología , Masculino , Fenoles/administración & dosificación , Distribución Aleatoria , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Accidente Cerebrovascular/prevención & control , Vasodilatación/efectos de los fármacos

RESUMEN

Incubation of aortic rings in a culture medium produces phenomena similar to those observed with aging, i.e. oxidative stress and inflammation leading to increased nitric oxide (NO)-mediated dilation and decreased arterial sensitivity to vasoconstrictor agents. We evaluated whether melatonin protects aortic rings from such a decrease in vasoreactivity. Two concentrations of melatonin were used: 10(-8) M, EC50 for vascular MT1-MT2 receptors, and 10(-5) M, reported as anti-oxidant. Anti-oxidant capacity, inducible nitric oxide synthase (iNOS) expression and isometric contraction of thoracic aorta rings (Wistar rats) evoked by norepinephrine (NE) were assessed. Three days of incubation of aortic rings induced iNOS expression and a fall in NE-evoked contraction. When melatonin was added to the organ bath, it (10(-5) M) increased (+96%, P < 0.05), but did not restore (compared with freshly isolated rings) NE-evoked contraction. Three days of treatment with melatonin increased (10(-8) M, +99%) or restored (10(-5) M, +216%) NE-evoked contraction (compared with freshly isolated rings). The beneficial effects of 10(-8) and 10(-5) M melatonin on NE-evoked contraction were abolished in the presence of luzindole (2 x 10(-6) M, a melatonin receptor antagonist). The incubation-induced increase in iNOS expression was reduced following 3 days of melatonin administration (10(-8) and 10(-5) M). Melatonin (10(-5) M) increased catalase activity (6550 +/- 256, P < 0.05 vs. nontreated fresh aortic rings 5554 +/- 444 nmol min(-1) mg protein(-1)). In conclusion, melatonin counteracts the incubation-induced loss of agonist-evoked contraction of aortic rings by a specific receptor-mediated phenomenon involving iNOS expression; at higher melatonin concentrations, an anti-oxidant effect is probably also involved.

Asunto(s)

Aorta Torácica/efectos de los fármacos , Melatonina/farmacología , Animales , Aorta Torácica/fisiología , Catalasa/metabolismo , Medios de Cultivo/farmacología , Glutatión Peroxidasa/metabolismo , Técnicas In Vitro , Masculino , Óxido Nítrico Sintasa de Tipo II/metabolismo , Norepinefrina/farmacología , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Vasoconstrictores/farmacología

RESUMEN

Two incidents of partner violence are investigated using qualitative methodology to discover strategies women use to protect themselves and examine women's use of violence. Data were collected from 447 women (age 18 or older) from 7 domestic violence programs and 5 substance use disorder treatment programs in a midwestern state. Women were found to have developed numerous self-protection strategies, some using nonphysical means only, others using physical means only, and others combining nonphysical and physical means. Women often used a variety of strategies in the same incident. Few women initiated violence against partners. Implications for theory and research are discussed.

Asunto(s)

Adaptación Psicológica , Mujeres Maltratadas/psicología , Autocuidado , Maltrato Conyugal/prevención & control , Adolescente , Adulto , Agresión/psicología , Reacción de Fuga , Femenino , Humanos , Medio Oeste de Estados Unidos , Motivación , Negociación/psicología , Comunicación Persuasiva , Investigación Cualitativa , Conducta Fugitiva/psicología , Autocuidado/métodos , Autocuidado/psicología , Maltrato Conyugal/psicología , Esposos/psicología , Trastornos Relacionados con Sustancias/psicología , Encuestas y Cuestionarios , Conducta Verbal

RESUMEN

The intracellular mechanism by which sepsis lowers vascular reactivity and the subsequent reversal by dexamethasone or nitric oxide synthase (NOS) inhibitors remain unclear. We measured the sensitivity of contraction of the rat tail artery to intracellular Ca2+ in a model of polymicrobial septic shock. At 22 h after cecal ligation and puncture (CLP), rats were treated with an anti-inflammatory glucocorticoid (dexamethasone, 1 mg/kg ip), an inducible NOS inhibitor (L-canavanine, 100 mg/kg ip), or saline. At 24 h after CLP, endothelium-denuded, perfused segments of tail artery were loaded with the intracellular Ca2+-sensitive dye fura 2 in vitro. Intracellular Ca2+ concentration and perfusion pressure were measured simultaneously. The rightward shift of the perfusion pressure-intracellular Ca2+ mobilization curve after norepinephrine stimulation subsequent to CLP indicates decreased intracellular Ca2+ sensitivity of contraction. The relation was restored by dexamethasone (which also restored in vivo blood pressure and flow), but not by L-canavanine (which restored perfusion pressure by further mobilization of intracellular Ca2+). We conclude that CLP lowers vasomotion by lowering intracellular Ca2+ sensitivity, which can be restored with glucocorticoid treatment. The involvement of inducible NOS does not solely account for the sepsis-induced reduction in Ca2+ sensitivity of contraction.

Asunto(s)

Calcio/fisiología , Canavanina/farmacología , Dexametasona/farmacología , Glucocorticoides/farmacología , Choque Séptico/fisiopatología , Vasoconstricción/efectos de los fármacos , Animales , Arginina/análogos & derivados , Arterias/efectos de los fármacos , Arterias/fisiología , Hemodinámica/efectos de los fármacos , Hemodinámica/fisiología , Masculino , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Ratas , Ratas Wistar , Cola (estructura animal)/irrigación sanguínea , Vasoconstricción/fisiología

RESUMEN

We studied the effect of age on the response of aortic rings to injury produced by three days' incubation, and the mechanism of this response. Five-mm rings of the thoracic aorta isolated from Wistar rats were incubated or not in culture medium. Isometric contraction evoked by agonists (norepinephrine or serotonin) or high [K(+)](e) was determined in the presence and absence of endothelium. Experiments were repeated in the presence of propranolol (0.3 microM), polymixin B (36 microM), pyrrolidine dithiocarbamate (50 microM) or glutathione (3 mM). Inductible NO-synthase and cyclo-oxygenase-2 mRNA were determined by real-time PCR, and glutathione-related enzymes and catalase activity by spectrophotometry. Incubation reduced the isometric contraction evoked by agonists but not by high [K(+)](e). The reduction in agonist-evoked contraction was greater in rings from adult (norepinephrine Emax-80%) than in young (-40%) rats. The removal of the endothelium had no effect. The reduction in norepinephrine-evoked contraction was not due to endotoxin contamination, beta-adrenoceptor-mediated dilation or any change in ring structure (no fibrosis or edema). Inductible NO-synthase (but not cyclo-oxygenase-2) mRNA increased on incubation. N(G)-nitro-L-arginine methyl ester partially restored contractility in rings from adult animals, further addition of an anti-oxidant restored norepinephrine-evoked contraction. Catalase fell with age and glutathione reductase increased upon incubation in rings from young donors only. In conclusion, incubation of the aorta produces a specific reduction in agonist-evoked contraction that involves induction of smooth muscle cell oxidative stress and iNOS. The reaction is greater in rings from older animals.

Asunto(s)

Aorta Torácica/fisiología , Medios de Cultivo/farmacología , Contracción Muscular/efectos de los fármacos , Músculo Liso Vascular/efectos de los fármacos , ARN Mensajero/metabolismo , Factores de Edad , Análisis de Varianza , Animales , Catalasa/metabolismo , Ciclooxigenasa 2 , Cartilla de ADN , Glutatión/farmacología , Isoenzimas/genética , Isoenzimas/metabolismo , Masculino , Contracción Muscular/fisiología , Músculo Liso Vascular/fisiología , Óxido Nítrico Sintasa/genética , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico Sintasa de Tipo II , Norepinefrina/farmacología , Potasio/farmacología , Propranolol/farmacología , Prostaglandina-Endoperóxido Sintasas/genética , Prostaglandina-Endoperóxido Sintasas/metabolismo , Pirrolidinas/farmacología , Ratas , Ratas Wistar , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Serotonina/farmacología , Espectrofotometría , Tiocarbamatos/farmacología , Factores de Tiempo