The Experts below are selected from a list of 393 Experts worldwide ranked by ideXlab platform

Manuel Ramírez - One of the best experts on this subject based on the ideXlab platform.

  • Brain Aminopeptidases and hypertension.
    Journal of the renin-angiotensin-aldosterone system : JRAAS, 2006
    Co-Authors: Inmaculada Banegas, Ana Belén Segarra, Isabel Prieto, Francisco Alba, Raquel Duran, Francisco Vives, Marc De Gasparo, Francisco Hermoso, Manuel Ramírez
    Abstract:

    The brain Aminopeptidases that participate in the enzymatic cascade of the renin-angiotensin system play a major role in blood pressure (BP) control, and their study offers new perspectives for the understanding of central BP control and the treatment of hypertension. In this system, angiotensin II is converted to angiotensin III (Ang III) by Glutamyl Aminopeptidase (GluAP) and Ang III is further metabolised to angiotensin IV by alanyl Aminopeptidase or arginine-Aminopeptidase. It is now clear that Ang III is the key active form of the central angiotensins, exerting tonic stimulatory control over BP. Therefore, the development of GluAP inhibitors as potential antihypertensive agents offers new perspectives for therapy. Brain aspartyl Aminopeptidase, which converts angiotensin I to angiotensin 2-10, is also a possible target for antihypertensive therapy because of its potential role in BP control. Finally, since changes in BP levels, that paralleled changes in brain and plasma Aminopeptidase activities, were observed after unilateral lesions of the nigrostriatal system, brain asymmetry, Aminopeptidase activities and BP control appear to be related, resulting their interplay in an asymmetrical neuroendocrine response that differentially affect BP control. The study of this interaction may contribute to our understanding of how the brain controls BP.

  • Ethanol modifies differently aspartyl- and Glutamyl-Aminopeptidase activities in mouse frontal cortex synaptosomes
    Brain research bulletin, 2002
    Co-Authors: María Dolores Mayas, María Jesús Ramírez-expósito, María Jesús García, Manuel Ramírez, José Manuel Martínez-martos
    Abstract:

    Aminopeptidase A activity (aspartyl Aminopeptidase (AspAP) and Glutamyl Aminopeptidase (GluAP) exerts angiotensinase activity due to its relation to the metabolism of angiotensins in the regional brain renin-angiotensin system (RAS). This activity may also modify the free amino acid pool through the release of N-terminal acidic amino acids. Ethanol (EtOH) exerts profound effects on the brain, inducing important neurological damages. Our purpose is to study the influence of EtOH on AspAP and GluAP activities on basal and K(+)-stimulated conditions, at the synapse level. We used mouse frontal cortex synaptosomes and their incubation supernatant in a Ca(2+)-containing or Ca(2+)-free artificial cerebrospinal fluid. We evaluate the possible contribution of these enzymatic activities on brain blood pressure regulation through RAS and/or the free acidic amino acid pool. The results obtained are correlated with several parameters of oxidative stress, such as free radical generation, lipid peroxidation, and protein oxidation. Under basal conditions, in synaptosomes, EtOH inhibits AspAP and GluAP activities independently of Ca(2+). In the supernatant, however, EtOH differently modulates the two enzyme activities under the various concentrations. Under K(+)-stimulated conditions, EtOH inhibits the K(+)-stimulated increase on AspAP and GluAP differently depending on the presence or absence of Ca(2+) and the concentration of EtOH used. These results invalidate the idea that excess free acidic amino acids could be released by AspAP and GluAP to induce neurodegeneration. The changes in AspAP and GluAP activities as a consequence of EtOH administration and their role in the brain RAS are discussed.

  • Serum Aminopeptidase A Activity of Mice Is Related to Dietary Fat Saturation
    The Journal of nutrition, 2001
    Co-Authors: G. Arechaga, José Antonio Muela Martínez, M. Ramirez, Isabel Prieto, Francisco Alba, Marc De Gasparo, María Jesús Fernández Sánchez, Manuel Ramírez
    Abstract:

    A high intake of monounsaturated fat has been proposed to be a dietary factor that can decrease the incidence of cardiovascular disease and hypertension. In addition, increasing dietary fat saturation has been shown to increase plasma total cholesterol and elevate systolic and diastolic blood pressures. We demonstrated previously that cholesterol selectively increases in vitro Aminopeptidase A activity, which is related to angiotensin metabolism. In this study, we investigated the effect of different degrees of dietary fatty acid saturation on serum Aminopeptidase activities in vivo. Serum total cholesterol concentrations were also measured. Five groups of male Balb/C mice were fed for 10 wk diets containing 2.4 g/100 g of sunflower oil, fish oil, olive oil, lard or coconut oil. We measured alanyl-, arginyl-, cystinyl-, pyroGlutamyl-, aspartyl- and Glutamyl-specific Aminopeptidase activities using arylamides as substrates. Serum total cholesterol levels were higher in mice fed diets containing saturated oils (lard and coconut) than in those consuming sunflower oil, which is unsaturated. Two of the serum Aminopeptidase A activities (aspartyl and Glutamyl Aminopeptidase) increased progressively with the degree of saturation of the dietary fatty acids; activities were significantly greater in mice fed coconut oil than in those fed sunflower or fish oil. Therefore, the substrates hydrolyzed by this activity as well as their functions may be similarly affected. These results may have some implication for the treatment of cardiovascular disease.

  • Comparative distribution of Glutamyl and aspartyl Aminopeptidase activities in mouse organs.
    Hormone and Metabolic Research, 2000
    Co-Authors: María Jesús Ramírez-expósito, Isabel Prieto, Francisco Alba, Manuel Ramírez
    Abstract:

    _ To evaluate the functional role of Glutamyl and aspartyl Aminopeptidases, their soluble and membrane-bound activities were measured simultaneously in several tissues of normal mice using arylamide derivatives as substrates. Although the soluble aspartyl Aminopeptidase activity showed its highest levels in the testicle, the rest of the activities presented their highest levels in the kidney. Different patterns of distribution were observed for Glutamyl and aspartyl Aminopeptidase activities and also for soluble and membrane-bound aspartyl Aminopeptidase activities. However no major differences were observed between soluble and membrane-bound Glutamyl Aminopeptidase activities. This unequal distribution suggests that the use of arylamide derivatives as substrates is a sensitive method that distinguishes between these enzymatic activities. The results also suggest different functions for soluble and membrane-bound aspartyl Aminopeptidase activities, and for Glutamyl and aspartyl Aminopeptidase activities.

  • Sex Differences and In Vitro Effects of Steroids on Serum Aminopeptidase Activities
    Peptides, 1998
    Co-Authors: José Antonio Muela Martínez, M. Ramirez, Isabel Prieto, Francisco Alba, Manuel Ramírez
    Abstract:

    Abstract We studied the possible existence of physiological sex differences in serum Aminopeptidase activities in mice, by evaluating the effect of gonadectomy and the in vitro response to the presence in the medium of cholesterol or steroid hormones. Alanyl- and Glutamyl-Aminopeptidase activities were measured in sera from male, female, orchiectomized and ovariectomized mice, incubated with substrate solutions, and compared with the same groups of serum incubated with substrate solutions including cholesterol, 17-β-estradiol, testosterone, progesterone or hydrocortisone. Our results demonstrated highly significant sex differences, and an influence of cholesterol and steroid hormones on Aminopeptidase activity. Depending on the nature of the Aminopeptidase, these enzymes responded in different ways to the presence of these substances and also responded differently to gonadectomy. For alanyl-Aminopeptidase activity, but not for Glutamyl-Aminopeptidase activity, there was a clear difference in response between males and females to incubation of the serum with steroid hormones.

José Manuel Martínez-martos - One of the best experts on this subject based on the ideXlab platform.

  • Ovarian Renin—Angiotensin System-regulating Aminopeptidases Are Involved in Progesterone Overproduction in Rats with Mammary Tumours Induced by N-Methyl Nitrosourea
    Anticancer research, 2009
    Co-Authors: M. P. Carrera, María Dolores Mayas, María Jesús Ramírez-expósito, María Jesús García, José Manuel Martínez-martos
    Abstract:

    A local renin-angiotensin system (RAS) has been found in ovary. This ovarian RAS may regulate ovarian steroidogenesis. Ample studies show that the ovarian hormones estradiol (E2) and progesterone (P) are strongly implicated in the development of breast cancer. Materials and Methods: The aim of the present work was to elucidate if alterations in ovarian RAS, analyzed through their proteolytic regulatory enzymes Aminopeptidase A (APA), B (APB) and N (APN), could be responsible for an altered steroidogenesis in rats with mammary tumours induced by N-methyl nitrosourea (NMU). Results: We describe here a highly significant increase of serum P levels in NMU-treated rats, concomitantly with an increase in ovarian aspartyl and Glutamyl Aminopeptidase activities (named together as APA activity). Moreover, we did not find changes in APB or APN activities, suggesting an increased metabolism from Ang II to Ang III and a decreased catabolism of Ang III. Conclusion: The relationship between ovarian RAS and P overproduction in a rat model of mammary carcinogenesis indicates ovarian RAS as a new potential target in breast cancer therapy.

  • Renin-Angiotensin System-Regulating Aminopeptidase Activities Are Modified in the Pineal Gland of Rats with Breast Cancer Induced by N-Methyl-Nitrosourea
    Cancer investigation, 2006
    Co-Authors: M. P. Carrera, María Dolores Mayas, María Jesús Ramírez-expósito, María Jesús García, M. T. Valenzuela, Basilio Dueñas, José Manuel Martínez-martos
    Abstract:

    Objective: Pineal function has been considered particularly as a neuroendocrine modulator in hormone responsive tumors, like the hormone-dependent mammary tumors. The complexity of the gland function, moreover, is denoted by the presence of a local renin-angiotensin-system (RAS) that regulates melatonin biosynthesis. Classically, angiotensin II (Ang II) has been considered as the effector peptide of the RAS, but Ang II is not the only active peptide. Several of its degradation products, including angiotensin III (Ang III) and angiotensin IV (Ang IV) also possess biological functions. These peptides are formed via the activity of several Aminopeptidases. Our aim is to know their role in the regulation of pineal RAS and breast cancer. Design: Aminopeptidase N (APN), Aminopeptidase B (APB) and Aminopeptidase A (aspartyl- and Glutamyl-Aminopeptidase, APA) activities are measured in the pineal gland of rats with breast cancer induced by N-methyl nitrosourea (NMU). Methods: Aminopeptidase activities were measur...

  • Glutamyl- but not aspartyl-Aminopeptidase activity is modified in serum of N-methyl nitrosourea-induced rat mammary tumours.
    Anticancer research, 2004
    Co-Authors: M. P. Carrera, María Dolores Mayas, María Jesús Ramírez-expósito, María Jesús García, M. T. Valenzuela, José Manuel Martínez-martos
    Abstract:

    Background: The rat model of breast cancer induced by the administration of N-methyl-nitrosourea (NMU) constitutes a useful tool for dissecting the initiation, promotion and progression process of carcinogenesis. Angiogenesis, the recruitment of new blood vessels, is an essential component of the metastatic pathway. Tumour vessels have an aberrant response to constrictor hormones, such as angiotensin II (Ang II). Ang II degradation to form angiotensin III (Ang III) begins with the action of Glutamyl Aminopeptidase (GluAP) and aspartyl Aminopeptidase (AspAP), named together as Aminopeptidase A activity (APA). The present work analyses GluAP and AspAP activities in serum of NMU-induced rat mammary tumours, to evaluate the putative value of these activities as biological markers of the initiation and promotion of the disease. Materials and Methods: Serum AspAP and GluAP activities were measured fluorimetrically using their corresponding aminoacyl-‚-naphthylamide. Results: The increase found in GluAP but not in AspAP suggests an increase in Ang III and a decrease in Ang II serum circulating levels. Conclusion: The decrease in Ang II may be responsible for the overexpression of AT1 receptors described in breast cancer. However, increased levels of Ang III, which exhibit the same affinity for the AT1 receptor, would favour the development of the disease.

  • Ethanol modifies differently aspartyl- and Glutamyl-Aminopeptidase activities in mouse frontal cortex synaptosomes
    Brain research bulletin, 2002
    Co-Authors: María Dolores Mayas, María Jesús Ramírez-expósito, María Jesús García, Manuel Ramírez, José Manuel Martínez-martos
    Abstract:

    Aminopeptidase A activity (aspartyl Aminopeptidase (AspAP) and Glutamyl Aminopeptidase (GluAP) exerts angiotensinase activity due to its relation to the metabolism of angiotensins in the regional brain renin-angiotensin system (RAS). This activity may also modify the free amino acid pool through the release of N-terminal acidic amino acids. Ethanol (EtOH) exerts profound effects on the brain, inducing important neurological damages. Our purpose is to study the influence of EtOH on AspAP and GluAP activities on basal and K(+)-stimulated conditions, at the synapse level. We used mouse frontal cortex synaptosomes and their incubation supernatant in a Ca(2+)-containing or Ca(2+)-free artificial cerebrospinal fluid. We evaluate the possible contribution of these enzymatic activities on brain blood pressure regulation through RAS and/or the free acidic amino acid pool. The results obtained are correlated with several parameters of oxidative stress, such as free radical generation, lipid peroxidation, and protein oxidation. Under basal conditions, in synaptosomes, EtOH inhibits AspAP and GluAP activities independently of Ca(2+). In the supernatant, however, EtOH differently modulates the two enzyme activities under the various concentrations. Under K(+)-stimulated conditions, EtOH inhibits the K(+)-stimulated increase on AspAP and GluAP differently depending on the presence or absence of Ca(2+) and the concentration of EtOH used. These results invalidate the idea that excess free acidic amino acids could be released by AspAP and GluAP to induce neurodegeneration. The changes in AspAP and GluAP activities as a consequence of EtOH administration and their role in the brain RAS are discussed.

Kyeong Kyu Kim - One of the best experts on this subject based on the ideXlab platform.

  • Structural and kinetic bases for the metal preference of the M18 Aminopeptidase from Pseudomonas aeruginosa.
    Biochemical and biophysical research communications, 2014
    Co-Authors: Duy Duc Nguyen, Doyoun Kim, Ramesh Pandian, Hye-jin Yoon, Kap Sun Kim, Kyung Hee Yun, Jin-hahn Kim, Kyeong Kyu Kim
    Abstract:

    Abstract The peptidases in clan MH are known as cocatalytic zinc peptidases that have two zinc ions in the active site, but their metal preference has not been rigorously investigated. In this study, the molecular basis for metal preference is provided from the structural and biochemical analyses. Kinetic studies of Pseudomonas aeruginosa aspartyl Aminopeptidase (PaAP) which belongs to peptidase family M18 in clan MH revealed that its peptidase activity is dependent on Co 2+ rather than Zn 2+ : the k cat (s −1 ) values of PaAP were 0.006, 5.10 and 0.43 in no-metal, Co 2+ , and Zn 2+  conditions, respectively. Consistently, addition of low concentrations of Co 2+ to PaAP previously saturated with Zn 2+ greatly enhanced the enzymatic activity, suggesting that Co 2+  may be the physiologically relevant cocatalytic metal ion of PaAP. The crystal structures of PaAP complexes with Co 2+ or Zn 2+ commonly showed two metal ions in the active site coordinated with three conserved residues and a bicarbonate ion in a tetragonal geometry. However, Co 2+ - and Zn 2+ -bound structures showed no noticeable alterations relevant to differential effects of metal species, except the relative orientation of Glu-265, a general base in the active site. The characterization of mutant PaAP revealed that the first metal binding site is primarily responsible for metal preference. Similar to PaAP, Streptococcus pneumonia Glutamyl Aminopeptidase (SpGP), belonging to Aminopeptidase family M42 in clan MH, also showed requirement for Co 2+ for maximum activity. These results proposed that clan MH peptidases might be a cocatalytic cobalt peptidase rather than a zinc-dependent peptidase.

  • One-step immobilization and purification of his-tagged enzyme using poly(2-acetamidoacrylic acid) hydrogel
    Macromolecular Research, 2013
    Co-Authors: Kyeong Kyu Kim, Hyung Soon Park, Sun-gu Lee, Jang-oo Lee, Hyun-jong Paik
    Abstract:

    Ni^2+-Complexed poly(2-acetamidoacrylic acid) (PAAA) hydrogel support was developed for the one-step immobilization and purification of recombinant histidine-tagged Glutamyl Aminopeptidase (His-tagged GAP). Ni^2+-PAAA hydrogel was prepared from the polymerization of 2-acetamidoacrylic acid (AAA) and 2,2-[(1,4-dioxo-1,4-butanediyl) diamino] bis(2-propenoic acid) (DBDBPA) with potassium persulfate in dimethyl sulfoxide (DMSO), followed by Ni^2+complexation. His-tagged GAP was immobilized directly from the cell lysate onto the Ni^2+-PAAA hydrogel support and then purified. Catalytic activity of immobilized His-tagged GAP for the hydrolysis of alanyl para -nitroanilide revealed 90% conversion after 30 min of incubation, indicating sustained catalytic activity. The hydrogel-immobilized enzyme also exhibited enhanced thermal stability of sustained 70% activity after 1 h incubation at 60 °C, while the free enzyme activity was reduced to 50% at the same condition. After four cycles of hydrogel regeneration, the immobilized enzyme lost only 20% of its initial activity. Ni^2+-PAAA hydrogel provided a new and convenient immobilization/purification system for His-tag enzymes through easy and simple procedures.

  • One-step immobilization and purification of his-tagged enzyme using poly(2-acetamidoacrylic acid) hydrogel
    Macromolecular Research, 2012
    Co-Authors: Kyeong Kyu Kim, Hyung Soon Park, Sun-gu Lee, Jang-oo Lee, Hyun-jong Paik
    Abstract:

    Ni2+-Complexed poly(2-acetamidoacrylic acid) (PAAA) hydrogel support was developed for the one-step immobilization and purification of recombinant histidine-tagged Glutamyl Aminopeptidase (His-tagged GAP). Ni2+-PAAA hydrogel was prepared from the polymerization of 2-acetamidoacrylic acid (AAA) and 2,2-[(1,4-dioxo-1,4-butanediyl) diamino] bis(2-propenoic acid) (DBDBPA) with potassium persulfate in dimethyl sulfoxide (DMSO), followed by Ni2+complexation. His-tagged GAP was immobilized directly from the cell lysate onto the Ni2+-PAAA hydrogel support and then purified. Catalytic activity of immobilized His-tagged GAP for the hydrolysis of alanylpara-nitroanilide revealed 90% conversion after 30 min of incubation, indicating sustained catalytic activity. The hydrogel-immobilized enzyme also exhibited enhanced thermal stability of sustained 70% activity after 1 h incubation at 60 °C, while the free enzyme activity was reduced to 50% at the same condition. After four cycles of hydrogel regeneration, the immobilized enzyme lost only 20% of its initial activity. Ni2+-PAAA hydrogel provided a new and convenient immobilization/purification system for His-tag enzymes through easy and simple procedures. Open image in new window

  • structural basis for the substrate specificity of pepa from streptococcus pneumoniae a dodecameric tetrahedral protease
    Biochemical and Biophysical Research Communications, 2010
    Co-Authors: Doyoun Kim, Boi Hoa San, Sang Hyun Moh, Hyejin Park, Dong Young Kim, Sangho Lee, Kyeong Kyu Kim
    Abstract:

    Regulated cytosolic proteolysis is one of the key cellular processes ensuring proper functioning of a cell. M42 family proteases show a broad spectrum of substrate specificities, but the structural basis for such diversity of the substrate specificities is lagging behind biochemical data. Here we report the crystal structure of PepA from Streptococcus pneumoniae, a Glutamyl Aminopeptidase belonging to M42 family (SpPepA). We found that Arg-257 in the substrate binding pocket is strategically positioned so that Arg-257 can make electrostatic interactions with the acidic residue of a substrate at its N-terminus. Structural comparison of the substrate binding pocket of the M42 family proteases, along with the structure-based multiple sequence alignment, argues that the appropriate electrostatic interactions contribute to the selective substrate specificity of SpPepA.

María Jesús Ramírez-expósito - One of the best experts on this subject based on the ideXlab platform.

  • Ovarian Renin—Angiotensin System-regulating Aminopeptidases Are Involved in Progesterone Overproduction in Rats with Mammary Tumours Induced by N-Methyl Nitrosourea
    Anticancer research, 2009
    Co-Authors: M. P. Carrera, María Dolores Mayas, María Jesús Ramírez-expósito, María Jesús García, José Manuel Martínez-martos
    Abstract:

    A local renin-angiotensin system (RAS) has been found in ovary. This ovarian RAS may regulate ovarian steroidogenesis. Ample studies show that the ovarian hormones estradiol (E2) and progesterone (P) are strongly implicated in the development of breast cancer. Materials and Methods: The aim of the present work was to elucidate if alterations in ovarian RAS, analyzed through their proteolytic regulatory enzymes Aminopeptidase A (APA), B (APB) and N (APN), could be responsible for an altered steroidogenesis in rats with mammary tumours induced by N-methyl nitrosourea (NMU). Results: We describe here a highly significant increase of serum P levels in NMU-treated rats, concomitantly with an increase in ovarian aspartyl and Glutamyl Aminopeptidase activities (named together as APA activity). Moreover, we did not find changes in APB or APN activities, suggesting an increased metabolism from Ang II to Ang III and a decreased catabolism of Ang III. Conclusion: The relationship between ovarian RAS and P overproduction in a rat model of mammary carcinogenesis indicates ovarian RAS as a new potential target in breast cancer therapy.

  • Renin-Angiotensin System-Regulating Aminopeptidase Activities Are Modified in the Pineal Gland of Rats with Breast Cancer Induced by N-Methyl-Nitrosourea
    Cancer investigation, 2006
    Co-Authors: M. P. Carrera, María Dolores Mayas, María Jesús Ramírez-expósito, María Jesús García, M. T. Valenzuela, Basilio Dueñas, José Manuel Martínez-martos
    Abstract:

    Objective: Pineal function has been considered particularly as a neuroendocrine modulator in hormone responsive tumors, like the hormone-dependent mammary tumors. The complexity of the gland function, moreover, is denoted by the presence of a local renin-angiotensin-system (RAS) that regulates melatonin biosynthesis. Classically, angiotensin II (Ang II) has been considered as the effector peptide of the RAS, but Ang II is not the only active peptide. Several of its degradation products, including angiotensin III (Ang III) and angiotensin IV (Ang IV) also possess biological functions. These peptides are formed via the activity of several Aminopeptidases. Our aim is to know their role in the regulation of pineal RAS and breast cancer. Design: Aminopeptidase N (APN), Aminopeptidase B (APB) and Aminopeptidase A (aspartyl- and Glutamyl-Aminopeptidase, APA) activities are measured in the pineal gland of rats with breast cancer induced by N-methyl nitrosourea (NMU). Methods: Aminopeptidase activities were measur...

  • Glutamyl- but not aspartyl-Aminopeptidase activity is modified in serum of N-methyl nitrosourea-induced rat mammary tumours.
    Anticancer research, 2004
    Co-Authors: M. P. Carrera, María Dolores Mayas, María Jesús Ramírez-expósito, María Jesús García, M. T. Valenzuela, José Manuel Martínez-martos
    Abstract:

    Background: The rat model of breast cancer induced by the administration of N-methyl-nitrosourea (NMU) constitutes a useful tool for dissecting the initiation, promotion and progression process of carcinogenesis. Angiogenesis, the recruitment of new blood vessels, is an essential component of the metastatic pathway. Tumour vessels have an aberrant response to constrictor hormones, such as angiotensin II (Ang II). Ang II degradation to form angiotensin III (Ang III) begins with the action of Glutamyl Aminopeptidase (GluAP) and aspartyl Aminopeptidase (AspAP), named together as Aminopeptidase A activity (APA). The present work analyses GluAP and AspAP activities in serum of NMU-induced rat mammary tumours, to evaluate the putative value of these activities as biological markers of the initiation and promotion of the disease. Materials and Methods: Serum AspAP and GluAP activities were measured fluorimetrically using their corresponding aminoacyl-‚-naphthylamide. Results: The increase found in GluAP but not in AspAP suggests an increase in Ang III and a decrease in Ang II serum circulating levels. Conclusion: The decrease in Ang II may be responsible for the overexpression of AT1 receptors described in breast cancer. However, increased levels of Ang III, which exhibit the same affinity for the AT1 receptor, would favour the development of the disease.

  • Ethanol modifies differently aspartyl- and Glutamyl-Aminopeptidase activities in mouse frontal cortex synaptosomes
    Brain research bulletin, 2002
    Co-Authors: María Dolores Mayas, María Jesús Ramírez-expósito, María Jesús García, Manuel Ramírez, José Manuel Martínez-martos
    Abstract:

    Aminopeptidase A activity (aspartyl Aminopeptidase (AspAP) and Glutamyl Aminopeptidase (GluAP) exerts angiotensinase activity due to its relation to the metabolism of angiotensins in the regional brain renin-angiotensin system (RAS). This activity may also modify the free amino acid pool through the release of N-terminal acidic amino acids. Ethanol (EtOH) exerts profound effects on the brain, inducing important neurological damages. Our purpose is to study the influence of EtOH on AspAP and GluAP activities on basal and K(+)-stimulated conditions, at the synapse level. We used mouse frontal cortex synaptosomes and their incubation supernatant in a Ca(2+)-containing or Ca(2+)-free artificial cerebrospinal fluid. We evaluate the possible contribution of these enzymatic activities on brain blood pressure regulation through RAS and/or the free acidic amino acid pool. The results obtained are correlated with several parameters of oxidative stress, such as free radical generation, lipid peroxidation, and protein oxidation. Under basal conditions, in synaptosomes, EtOH inhibits AspAP and GluAP activities independently of Ca(2+). In the supernatant, however, EtOH differently modulates the two enzyme activities under the various concentrations. Under K(+)-stimulated conditions, EtOH inhibits the K(+)-stimulated increase on AspAP and GluAP differently depending on the presence or absence of Ca(2+) and the concentration of EtOH used. These results invalidate the idea that excess free acidic amino acids could be released by AspAP and GluAP to induce neurodegeneration. The changes in AspAP and GluAP activities as a consequence of EtOH administration and their role in the brain RAS are discussed.

  • Comparative distribution of Glutamyl and aspartyl Aminopeptidase activities in mouse organs.
    Hormone and Metabolic Research, 2000
    Co-Authors: María Jesús Ramírez-expósito, Isabel Prieto, Francisco Alba, Manuel Ramírez
    Abstract:

    _ To evaluate the functional role of Glutamyl and aspartyl Aminopeptidases, their soluble and membrane-bound activities were measured simultaneously in several tissues of normal mice using arylamide derivatives as substrates. Although the soluble aspartyl Aminopeptidase activity showed its highest levels in the testicle, the rest of the activities presented their highest levels in the kidney. Different patterns of distribution were observed for Glutamyl and aspartyl Aminopeptidase activities and also for soluble and membrane-bound aspartyl Aminopeptidase activities. However no major differences were observed between soluble and membrane-bound Glutamyl Aminopeptidase activities. This unequal distribution suggests that the use of arylamide derivatives as substrates is a sensitive method that distinguishes between these enzymatic activities. The results also suggest different functions for soluble and membrane-bound aspartyl Aminopeptidase activities, and for Glutamyl and aspartyl Aminopeptidase activities.

Rosemary Wangensteen - One of the best experts on this subject based on the ideXlab platform.

  • Image_1_Klotho and Aminopeptidases as Early Biomarkers of Renal Injury in Zucker Obese Rats.tif
    2018
    Co-Authors: Sebastián Montoro-molina, Andrés Quesada, Félix Vargas, Antonio Osuna, Francisco O’valle, Antonio López-carmona, Natividad Martín-morales, Rosemary Wangensteen
    Abstract:

    The aim of this study was to investigate if urinary Glutamyl Aminopeptidase (GluAp), alanyl Aminopeptidase (AlaAp), Klotho and hydroxyproline can be considered as potential biomarkers of renal injury and fibrosis in an experimental model of obesity. Male Zucker lean (ZL) and obese (ZO) rats were studied from 2 to 8 months old. Kidneys from ZO rats at the end of the study (8 months old) developed mild focal and segmental glomerulosclerosis as well as moderate tubulointerstitial injury. Urinary excretion of Klotho was higher in ZO rats at 2, 5, and 8 months of study, plasma Klotho levels were reduced and protein abundance of Klotho in renal tissue was similar in ZL and ZO rats. GluAp and AlaAp urinary activities were also increased in ZO rats throughout the time-course study. ZO rats showed an augmentation of hydroxyproline content in renal tissue and a significant increase of tubulointerstitial fibrosis. Correlation studies demonstrated that GluAp, AlaAp, and Klotho are early diagnostic markers of renal lesions in Zucker obese rats. Proteinuria and hydroxyproline can be considered delayed diagnostic markers because their contribution to diagnosis starts later. Another relevant result is that GluAp, AlaAp, and Klotho are related not only with diagnosis but also with prognosis of renal lesions in Zucker obese rats. Moreover, strong predictive correlations of aminopeptidasic activities with the percentage of renal fibrosis or with renal hydroxyproline content at the end of the experiment were observed, indicating that an early increased excretion of these markers is related with a higher later extent of fibrosis in Zucker obese rats. In conclusion, GluAp, AlaAp, and Klotho are early diagnostic markers that are also related with the extent of renal fibrosis in Zucker obese rats. Therefore, they have a potential use not only in diagnosis, but also in prognosis of obesity-associated renal lesions.

  • Glutamyl Aminopeptidase in microvesicular and exosomal fractions of urine is related with renal dysfunction in cisplatin-treated rats
    2017
    Co-Authors: Andrés Quesada, Sebastián Montoro-molina, Félix Vargas, María Del Carmen De Gracia, Antonio Osuna, Ana Belén Segarra, Manuel Gómez-guzmán, Francisco O’valle, Rosemary Wangensteen
    Abstract:

    PurposeThe aim of this work was to investigate if the content of Glutamyl Aminopeptidase (GluAp) in microvesicular and exosomal fractions of urine is related with renal dysfunction in cisplatin-treated rats.MethodsUrine samples were collected 24 hours after injection of cisplatin (7 mg/kg, n = 10) or saline serum (n = 10), and they were subjected to differential centrifugation at 1.000, 17.000 and 200.000 g to obtain microvesicular and exosomal fractions. GluAp was measured with a commercial ELISA kit in both fractions. Serum creatinine (SCr) and body weight were measured 15 days after treatment. We analyzed if early excretion of GluAp in microsomal and exosomal fractions was correlated with final SCr and body weight increase. In a second experiment, enzymatic activities of GluAp and alanyl Aminopeptidase (AlaAp) in urine, microvesicular and exosomal fractions were measured three days after injection. We analyzed the correlation of both markers with SCr determined at this point. Finally, we studied the expression of GluAp and extracellular vesicles markers Alix and tumor susceptibility gene (TSG101) in both fractions by immunoblotting.ResultsGluAp excretion was increased in all fractions of urine after cisplatin treatment, even if data were normalized per mg of creatinine, per body weight or per total protein content of each fraction. We found significant predictive correlations with SCr concentration, and inverse correlations with body weight increase determined 15 days later. Three days after injection, aminopeptidasic activities were markedly increased in all fractions of urine in cisplatin-treated rats. The highest correlation coefficient with SCr was found for GluAp in microvesicular fraction. Increase of GluAp in microvesicular and exosomal fractions from cisplatin-treated rats was confirmed by immunoblotting. Alix and TSG101 showed different patterns of expression in each fraction.ConclusionsDetermination of GluAp content or its enzymatic activity in microvesicular and exosomal fractions of urine is an early and predictive biomarker of renal dysfunction in cisplatin-induced nephrotoxicity. Measurement of GluAp in these fractions can serve to detect proximal tubular damage independently of glomerular filtration status.

  • Immunological detection of Glutamyl Aminopeptidase in urine samples from cisplatin-treated rats.
    Proteomics. Clinical applications, 2015
    Co-Authors: Sebastián Montoro-molina, Andrés Quesada, Piedad V. Zafra-ruiz, Francisco O'valle, Félix Vargas, María Del Carmen De Gracia, Antonio Osuna, Rosemary Wangensteen
    Abstract:

    Purpose The aim of this work is to demonstrate if urinary excretion of Glutamyl Aminopeptidase (GluAp) can be quantified by immunological methods. Experimental design Urine samples from control and cisplatin-treated rats (n = 10 each group) were obtained at 1, 8, and 15 days after cisplatin injection. GluAp was analyzed by kinetic fluorimetry, ELISA, and immunoblotting. Sensitivity and specificity was studied for fluorimetric activity and ELISA 24 h after cisplatin injection. We also analyzed the predictive value over renal dysfunction at the end of the experiment. Results GluAp was easily detected by immunoblotting and ELISA, and its urinary excretion was increased in cisplatin-treated rats (p < 0.01). Results obtained with ELISA were strongly correlated (r = 0.8186; p < 0.0001) with fluorimetric activity. Kinetic fluorimetry was the method with the highest AUC (AUC = 1) and the highest predictive value over serum creatinine (r = 0.7630; p = 0.0001) and body weight increase (r = –0.8721; p < 0.0001). Conclusions and clinical relevance GluAp can be detected in urine samples with immunological methods, making possible the development of an antibody-based kit for its determination. Its excretion correlates with the extent of renal dysfunction in cisplatin-treated rats, confirming its value as an early marker of renal damage that can be a diagnostic aid in renal diseases.

  • Glutamyl Aminopeptidase in microvesicular and exosomal fractions of urine is related with renal dysfunction in cisplatin-treated rats.
    Public Library of Science (PLoS), 1
    Co-Authors: Andrés Quesada, Sebastián Montoro-molina, Francisco O'valle, Félix Vargas, María Del Carmen De Gracia, Antonio Osuna, Ana Belén Segarra, Manuel Gómez-guzmán, Rosemary Wangensteen
    Abstract:

    The aim of this work was to investigate if the content of Glutamyl Aminopeptidase (GluAp) in microvesicular and exosomal fractions of urine is related with renal dysfunction in cisplatin-treated rats.Urine samples were collected 24 hours after injection of cisplatin (7 mg/kg, n = 10) or saline serum (n = 10), and they were subjected to differential centrifugation at 1.000, 17.000 and 200.000 g to obtain microvesicular and exosomal fractions. GluAp was measured with a commercial ELISA kit in both fractions. Serum creatinine (SCr) and body weight were measured 15 days after treatment. We analyzed if early excretion of GluAp in microsomal and exosomal fractions was correlated with final SCr and body weight increase. In a second experiment, enzymatic activities of GluAp and alanyl Aminopeptidase (AlaAp) in urine, microvesicular and exosomal fractions were measured three days after injection. We analyzed the correlation of both markers with SCr determined at this point. Finally, we studied the expression of GluAp and extracellular vesicles markers Alix and tumor susceptibility gene (TSG101) in both fractions by immunoblotting.GluAp excretion was increased in all fractions of urine after cisplatin treatment, even if data were normalized per mg of creatinine, per body weight or per total protein content of each fraction. We found significant predictive correlations with SCr concentration, and inverse correlations with body weight increase determined 15 days later. Three days after injection, aminopeptidasic activities were markedly increased in all fractions of urine in cisplatin-treated rats. The highest correlation coefficient with SCr was found for GluAp in microvesicular fraction. Increase of GluAp in microvesicular and exosomal fractions from cisplatin-treated rats was confirmed by immunoblotting. Alix and TSG101 showed different patterns of expression in each fraction.Determination of GluAp content or its enzymatic activity in microvesicular and exosomal fractions of urine is an early and predictive biomarker of renal dysfunction in cisplatin-induced nephrotoxicity. Measurement of GluAp in these fractions can serve to detect proximal tubular damage independently of glomerular filtration status